microbio final

lac operon

group of genes involved in lactose metabolism as cell uses lactose for growth (when no glucose) components: -the three genes: LacZ, LacY, LacA needed to make enzymes -promoter region is not match to sigma subunit so an activator (protein CAP) is needed to help bring sigma to bind to promoter -LacI: the repressor protein that is the on and off switch allowing organism to use lactose at needed times --> off when glucose present since glucose is a better E source and w/o lactose, it binds operator and prevents transcription -LacZ: encodes B-galactosidase, which breaks down/hydrolyzes lactose -LacY: encodes permease, which carries lactose across the membrane in the cell -LacA: encodes for AcT, which acetylates lactose as it is taken up by the cell

virus

not alive -did not evolve from LUCA, no common viral ancestor -derived from DNA of other cells -infectious, nonliving particles rely on cells for reproduction: -no ribosomes (use host ribosomes) -no metabolism as cannot make own ATP -no energy harvest -no energy storage -no retension of structure attaches to surface of cell and injects genetic material into cell, particles use cell's replication machinery and ribosomes to replicate and spread to other cells

parasites -obligate and facultative -intermediate, definitive and incidental host

organism that lives on or in a host and relies on it for growth and reproduction -obligate: requires host to live -facultative: host is optional, can survive outside host Sexual reproduction only within a host 3 types of hosts: -intermediate: no new offspring, can only spread -definitive: parasites create new offspring -incidental: parasites go to host accidentally as the host is not beneficial so no reproduction

What is PFU and how does it compare to CFU?

plaque-forming units: concentrations of viral suspensions measured by the number of plaques -host cells (bacteria or eukaryote) on plate, add viral particle, dead cells are the white spots, plaque -virus infects cells, killing them, creating plaques it is similar to CFU as bacteria is also placed on a plate but with just a medium and colonies also form, allowing for it to be counted but virus is counted by looking at number of dead cell plaques while bacteria is counted by looking at its own form

flagella polar vs many

polar flagellum: -cell with single polar flagellum moves right to left when flagella turns counterclockwise and tumbles when clockwise many flagella: -cell w many flagella all over its surface with counterclockwise rotation making coherent flagellar bundle and smooth movement -clockwise rotation is when bundle flies apart with tumbling resulting

bacteria colonization resistance

prevention of pathogen growth -antibiotics kill commensal microbes -clostridium difficile spore survive and cause disease -treatment: fecal transplant restores commensal microbes by introducing someone else's healthy microbes and cures disease -bad bc: diff btwn host and donor's microbes, which microbes, body rejects, delivery problems as O2 can kill beneficial microbes, invasive

Chemotaxis 2 component system

-CHE A is the attractant (sensor) and CHE Y is the repellant (response regulator) -no attractant, repellant or danger: CheA sensor autophosphorylates and triggers response with the P goes to Che Y which runs down the cell, leading to a change in flagellar rotation from default CCW to CW, causing the cell to tumble -return to normal by removing P not on when going towards attractant

Bacteroides

-G- -carbohydrate generalists -make SCFA -aerotolerant for short periods

What is an intermediate host vs a definitive host for a parasite? Distinguish between sexual reproduction and asexual reproduction

-Intermediate host is when the virus creates no new offspring, it can only spread and the host isn't harmed, it's just used for virus to get to definitive host -Definitive host is when parasites create new offspring in hist successfully -Sexual reproduction involves the fusion of male and female gametes (sperm and egg) to produce genetically diverse offspring with unique combinations of genes from both parents -Asexual reproduction, on the other hand, involves the production of offspring without the fusion of gametes, resulting in offspring that are genetically identical to the parent and to each other

Genetic drift and shift

-drift: point mutations = slow -shift: mutations causing reassortment , viruses infect same host cell = rapid

Explain how the lac operon is regulated, including the genes lacZ, lacY, lacA, and lacI. How is transcription controlled for each of these genes? You should understand how LacI binds to the operator (when does it bind, what does it not bind) and how CAP activates the lac operon in the absence of glucose. What would you expect in the following conditions? Why? 1. High glucose, high lactose 2. High glucose, no lactose 3. Low glucose, high lactose 4. Low glucose, no lactose

-The expression of the lac operon is regulated by the LacI protein, which binds to the operator sequence of the operon and inhibits transcription of the structural genes, and the amount of glucose present as glucose is the preferred energy source 1. High glucose, high lactose: -glucose represses CAP activator from binding, lactose prevents LacI from binding = no transcription 2. High glucose, no lactose -glucose binds CAP and LacI binds since no lactose preventing it = no transcription 3. Low glucose, high lactose -CAP binds activator site and lactose inhibits LacI from binding promoter = transcription 4. low glucose, no lactose -CAP binds and LacI binds to operator = no transcription Glucose present = no lactose made since its preferred

Malaria

-caused by a Apicomplexans -difinitive host is a mosquito, with parasite reproducing it in and spreading in it -mosquito w virus bites human (intermediate host where virus cannot reproduce) with virus going into bloodstream, then liver where it makes red blood cells rupture -symptom: red blood cell lysis = anemia

microbe species: community composition

-community composition : make up of a specific group of species that interact -16S rRNA gene sequencing: has constant conserved regions present in all species and variable regions that can be matched by primers and sequenced to find species type -metagenomic sequencing: technique used to study the genetic material from a complex mixture of microorganisms by extracting genetic material from a sample by breaking up bacteria and sequencing DNA fragments into genomes of various species present

stationary phase

-default condition for many microbes -cells look diff in this phase than they did when growing since their metabolism was altered to increase survival, not growth -nutrients depleted, can't find food freely = stressed -cells clumped together as stressor results in phenotype change -sRNA integrators process signals from diff stressors and integrate them into a single action and stimulates sigmaS (RpoS) synthesis, a regulator that turns on various genes on RpoS regulon -sigmaS regulates stationary phase response and all genes regulated by stationary phase -genes in stationary phase regulon induce multiple changes in cell (decrease cell vol, morphology change) -sigmaS competes w sigma70 to bind RNAP, growth-related functions turned off

kinetoplastids

-flagellate -lacks enzyme gyrase so all DNA in mitochondria -kinetoplast: network of DNA inside a large mitochondria -circles of DNA bound together with flagella moving it around 2 types of this insect parasite (leishmania major = sandfly) 1. trypanosoma cruzi = kissing big 2. trypanosoma brucei = tse tse fly -live extracellularly and survive in bloodstream as it doesn't go inside side -on skin causing lesions, kills skin cells and opens wounds

bacteria habitat - host intestine

-food broken down and absorbed in SI: sugars/starch (glucose), fat (lipids), protein (a.a) -dietary fiber doesn't get used, passes thru (plant carbohydrates) foregut fermenter: large stomachs containing bacteria that break down food (i.e monkey) Hindgut fermenter: large colon bc microbial fermentation occurs there, small stomachs (i.e humans)

no microbes in body (germ-free)

-huge cecum -less body fat even tho eating more but harvesting less E from diet -can't turn diet into nutrients since no mircrobes -vitamin deficiency, can die -skewed immune system -easily infected since no microbes to compete for nutrients with infectious microbes

Membrane proteins

-hydrophobic -70% bacterial membrane mass -how membrane proteins are embedded into membrane -signal peptide is on N terminus of target protein (15-30 a.a long) that is recognized and binds to signal recognition particle (SRP) -SRP grabs signal peptide and holds onto it to ground into channel or membrane -SRP escorts SS to dedicated secretory proteins in cell membrane with hydrophobic signal peptide part of protein inside membrane and hydrophilic part outside

Quorum sensing

-inducing a response when cells have reached a certain population number -the regulation of gene expression in response to fluctuations in cell-population density -bacterial communication based on cell density where bacteria senses other bacteria around them and a response activated when cell density at certain level -based on soluble signal in gram-positive bacteria: -an important mechanism for regulating gene expression and behavior in response to changes in population density, allowing bacteria to coordinate their actions and adapt to changes in their environment -a process of cell-to-cell communication that relies on the production and detection of small signaling molecules called autoinducing peptides (AIPs) -The sensor histidine kinase detects the presence of AIPs in the environment and phosphorylates the response regulator, which then activates or represses gene expression

Antiviral

-mimic host surface: tamiflu blocks NA protein and virion release blocked -block viral sensing: rimantadine: blocks M2 channel, blocks H+ sensing and invasion

grouping viruses - viral diversity

-no membrane, no helical capsid -membrane, no helical capsid -no membrane, helical capsid -membrane, helical capsid

colonization of a host by bacteria

-on outside surfaces majority of bacteria live in body cavities bc -resistance to physical disruption -nutrients provided and stored -is stable as bacteria lives off of host's actions to maintain homeostasis -reduces extremes of stress -doesn't trigger host's immune system -in tissues is bad bc it can kill host and immune system can kill bacteria

Explain the difference between a gene, operon, and regulon.

-operon: multiple genes under control of same promoter on mRNA strand --> each gene has own SD to make own protein -regulon: more than one operon that is under control of a single regulatory protein --> collection of genes that a sigma factor controls and promotes Gene: A region of DNA that codes for a specific protein

protozoa -amobae, apicomplexans, and flagellates

-protists, broad term, have lived long -dangerous microbes -amoebae: single-celled organism, usually harmless (pathogenic in brain tissue), moves like slug -apicomplexans and flagellates: contain parasites and need host to survive and replicate -infects and causes diseases in humans and don't respond to antibiotics -are not a single related group --> spread all over phylogenetic tree

Algae

-single-celled plant -photoautotrophs (E from sun, inorganic C source CO2) -cellulose cell wall makes it rigid -live in marine/aquatic environments -eukaryotic microbe

LuxI-LuxR system

-virbrio fischeri is a bacteria living in squid and is used to hide squid from predators in deep sea -enzyme LuxI makes signal/autoinducer AHL and response regulator Luxr that binds the signal -binding the autoinducer activates LuxR and induces expression of 7-gene Lux operon, encoding proteins that convert ATP into light -after bound by autoinducer, cell density decreases so light turns off -squid expels bacteria when done and density too high -light emission from squid depends on concentration of bacteria -low cell density: luxR operon off -high cell density: luxR turned on and genes on for bioluminescent

Fungi

-yeast -round-shaped, white -found in wild -chemoheterotrophs: degrade dead plant material to get C source -becomes infectious with filamentation, going from round to branch-like structure making hyphae -can be pathogenic and infect plants -sporulation: spread to new habitats and survive harsh environments -eukaryotic microbe

3 proteins in membrane stress

1. CPX P: senses misfolded proteins and will release CPXA to send stress signal and binds & blocks CPXA when not bound to misfolded protein 2. CPXA is a membrane-bound protein in periplasm that senses membrane stress, autophosphorylates on histidine kinase via ATP hydrolysis, and activates CPXR by giving it the phopshate 3. CPXR is the response regulator that takes P and activates transcription of genes needed Resolution occurs with the dephosphorylation of Cpx R

Name 3 fundamental processes that are conserved in all organisms

1. DNA replication: This process is essential for the transmission of genetic information from one generation to the next and is conserved in all organisms. 2. Protein synthesis: Proteins are essential for many biological processes, including metabolism, regulation, and defense. The basic steps of protein synthesis are conserved in all organisms, although the details may vary. 3. Energy metabolism: All organisms require energy to carry out their basic functions. Energy metabolism involves the conversion of nutrients into energy that can be used by the cell.

Name 3 fundamental qualities that are unique to eukaryotes (not found in any prokaryotes)

1. Membrane-bound organelles: Eukaryotic cells have a variety of specialized membrane-bound organelles, such as the mitochondria, endoplasmic reticulum, and Golgi apparatus. These organelles compartmentalize the cell and allow for the separation and specialization of different cellular functions. 2. Cytoskeleton: Eukaryotic cells possess a cytoskeleton that provides structural support for the cell and is involved in cell movement, division, and organization. 3. Sexual reproduction: Eukaryotic organisms reproduce sexually, involving the fusion of haploid gametes, allowing for the generation of genetic diversity in offspring and the evolution of new traits. Prokaryotic organisms, on the other hand, reproduce asexually by binary fission

All viruses follow the same general 5 steps when completing their life cycle (i.e. starting with one virus infecting a cell and ending up with multiple viruses getting released). What are these and what is happening at each step?

1. The binding and entry of the virus is when the receptors recognize the spike proteins, causing the virus to bind to cell 2. The genome is released into the cell after virus injects itself into host cell, with its capsid being destroyed 3. Early genes first expressed in virus creates genome replication proteins that take over the cell 4. Late genes are created, making virion proteins for structural reasons to make new viruses (makes capsid, spike proteins) 5. virion released from cell (not dead) by opening membrane and finds new cell to make more virus particles

viral life cycle (6 steps)

1. binding: receptors on host recognize spike proteins and virus binds to cell 2. virus enters: Once attached, the virus can then inject its genetic material, such as RNA or DNA, into the host cell 3. genome released into cell with outer protective layer capsid destroyed, virus degraded 4. early genes made with genome replication proteins made from first genes expressed in virus taking over cell 5. late genes: virion proteins made for structural reasons, new virus made (capsid, spike protein) 6. virion release, cell intact and finds new cell to make more virus particles

viral life cycle graph -4 steps

1. innoculation: amount of virions added (in a dish of cells) with low amounts of virions 2. eclipse: virus is disassembled so virus pop decreased after released into cell and destroyed 3. release: not synchronous, sharp increase in virion pop as virus is being made and released out of host cell 4. plateau: no more living hosts to infect, so stable, stationary virus pop

Graph of a viral replication curve: 1. What is each stage called and what is happening at each stage? 2. If I took the same graph, but told you it was a bacterial growth curve and changed the y axis to "# of bacteria," how would you interpret the data? The lines look similar, does that mean the same things are happening? Explain

1. innoculation: amount of virions added (in a dish of cells) with low amounts of virions 2. eclipse: virus is disassembled so virus pop decreased after released into cell and destroyed 3. release: not synchronous, sharp increase in virion pop as virus is being made and released out of host cell 4. plateau: no more living hosts to infect, so stable, stationary virus pop Number of bacteria would mean that this shows the growth of bacteria at different stages of growth, from when its abundant with nutrients available to nutrients being depleted and no more bacteria being made. -similar things are not happening as the virions are not growing and are introduced to the cells as is, where it then has a dip in the curve when it gets destroyed. We see a steady initial increase of the bacterial population with it growing with nutrients and then exponentially increasing, to it plateauing - Both bacteria and virions grow in the beginning and then have a stational phase with a plateau after their necessities have been depleted (nutrients, hosts), but the release phase is not synchronous like the bacteria's log phase.

Microbe benefits

1. regulation of our immune system 2. protection against other bacteria that cause disease 3. aid in food digestion 4. supply energy and vitamins

strategy for dealing with cell stress

1. sensing: happens outside bacteria and relayed into cell membrane, absolute vs dynamic 2. signal transduction: chain reaction with protein being modified 3. response: gene regulation increased or decreased, turn on series of genes to overcome stress 4. resolution: feedback inhibition with genes turned off

2 component systems -histidine kinase:

1. sensor: transmembrane protein partly inside and outside of cell, and cytoplasmic kinase with phosphorylation creating a change in protein to relay stressor 2. response regulator: activates transcription when phosphorylated -stressor comes to membrane protein -histidine kinase: senses changes in the external environment, triggering the autophosphorylation of hisitidine residue in sensory kinase (from ATP), which then phosphorylates the aspartic acid of response regulator. This activates the response regulator, which takes signal and binds control regions of target response genes to control their expression -feedback control with the dephosphorylation of the response regulator when stress response no longer needed so everything goes back to normal sensing in inner membrane, signal transduction in cytoplasm, response in nucleoid

other types of transcriptional regulation

4. transcription termination: Rho binds mRNA, transcripts are prematurely terminated after initiation 5. sRNA: decreases transcr. initiation and stops RNAP elongation, folds over and covers SD so that ribosomes are blocked but either blocks or exposes SD sequence to increase or decrease transl 6. mRNA stability: minor in bacteria, degradation of target mRNA molecules 7. translation: intiation can be repressed or activated, binding proteins to own mRNA prevents their translation 8. protein stability: sigma stabilized when bound tighly to RNAP and protected to proteolysis

What is a virus? What are 3 features present in all viruses (defining features)?

A virus is a small infectious agent that can only replicate inside the living cells of other organisms, 1. Capsid: protein shell that encloses genetic material (RNA or DNA) 2. Genome: DNA or RNA genetic info that makes host create more virus 3. Spike protein: can't sense the environment, binds to the cell's cell receptors

What would it mean if a viral "growth" curve is a flat line? Describe how this mightoccur

A flat curve is when the virion is in the plateau stage with no more living host cells to infect. This would occur when the virus has killed all of the host cells and can no longer increase its population since there are no more host cells to use for their machinery

flagella prok.

A long, whip-like filament that helps in cell motility -mainly in G- and requires energy H+ -rotates by turning flagellar filaments using energy from flow of protons from the transmembrane ion gradient -counterclockwise: flagella entwine into stiff bundles making a propeller-like force that results in desired motion = run -clockwise: bundles dissociated causing cell to tumble (stops and changes direction) in place -can alternate between both: brief runs interrupted by tumbling with new runs occurring in direction that cell faces after a tumble bacteria moves towards chemical attractant (i.e nutrient source) with longer-lasting runs in direction of chemical

Apicomplexans -micronemes, rhoptries, nucleus

A type of parasitic protozoan. -Some cause serious human disease -apical complex located at one end of the cell -secretory organelle with proteins secreted by vesicle by merging w membrane = membrane trafficking -micronemes: membrane-bound components that bind the host cell and invade host -rhoptries: proteins inside of it released after invasion and take control of host cells and their biology = host manipulation -nucleus: huge genome (80Mbp) ex. invaded mouse cells with parasites taking over cells, making mice unafraid of cats --> cats eat them and get parasite

Name two apicomplexan parasites and two kinetoplastid parasites. (Genus and species)

Apicomplexan parasites: 1. Plasmodium, the causative agent of malaria with mosquito host 2. Toxoplasma gondii, which can cause severe disease in humans and other animals, making mice not scared of cats Kinetoplastid parasites: insect parasite (leishmania major = sandfly), kills skin cells and opens wounds 1. trypanosoma cruzi = kissing big 2. trypanosoma brucei = tse tse fly

How do bacteria sense a rise in temperature? What is the sensor? What is the response regulator? How do they activate the σΗ regulon? How do they turn it off?

At high temperatures, the folded mRNA strands are melted, exposing the SD that allows for sigmaH to be made in high amounts, causing it to bind RNAP and have it turn on heat response genes on heat shock regulon -the sensor is sigmaH which increases affinity of RNAP for promoters of heat shock regulon and induces transcription of heat-shock related genes -the response regulator is DnaK ? which is a protein chaperon that facilitates proper protein folding SigmaH allows for RNAP to bind to heat shock regulon, producing DnaK which folds proteins. It is turned off by FTSH, a protease that degrades the bound DnaK and sigmaH and turns off system

Explain how CheA / CheY controls flagellar motion. How does this relate to a run vs tumble?Explain why the rotation direction of the flagella needs to be controlled.

CheA sensor gets signal, then autophosphorylates and triggers response with the P goes to Che Y which runs down the cell, leading to a change in flagellar rotation from default CCW to CW, causing cell to tumble -direction needs to be in control in order for bacteria to steer clear of danger as phosphorylating the CheA and then CheY causes a conformation change making the direction change to clockwise, resulting in a tumble to avoid the danger

What is chemotaxis? Can bacteria swim directly towards a chemoattractant?

Chemotaxis is the ability of bacteria to move towards or away from chemical gradients in their environment. Bacteria use chemotaxis to navigate towards nutrients or avoid toxins and other harmful compounds -Bacteria do not swim directly towards a chemoattractant, they do runs and tumbles, with runs being movements forward in a straight line for a short distance, and then tumbles when it stops and changes direction in a random manner -runs last longer in direction of attractant

biofilms

Colonies of bacteria that adhere together and adhere to environmental surfaces. -formation of organized microbial communities on surfaces -structure tailored to environment and has enhanced resistance to antibiotics and toxic substances -covers bacteria with extracellular matrix (waste materials made of polysaccharides, protein, DNA, lipids)

eukaryotic motility

flagella and cilia -flagella is a single structure while cilia coats the entire cell and moves in unison -made of microtubules -encased in membrane so cell uses ATP to power movement

Viral diversity DNA vs RNA genome

DNA genome: -less prone to degradation -low error rate bc of DNAP -replicate in host nucleus (host DNAP) -bigger genome (1Mb max) RNA genome: -susceptible to RNAse in host to degradation -higher mutation rate -stay in cytoplasm (viral RNAP) -sars cov2 (30kb) --> smaller viral genomes need to be replicated and be converted to mRNA using host DNAP, host RNAP, host ribosome

actinobacteria

G+ -bifidobacteria -milk carbohydrates -beneficial

fermicutes

G+ -degrades cellulose -aerosensitive -spores that can tolerate toxic O2

protein secretion

G+: thru cell wall via channel G-: thru inner and outer membranes via channels General systems: 1. membrane proteins: hydrophobicity 2. unfolded protein export: SEC system 3. folded protein export: TAT system 4. double membrane export: BAM system proteins with hydrophilic surface cannot readily enter hydrophobic phospholipid membrane

proteobacteria

G- -aerobes -many pathogens (i.e cholera) -lots present in newborns but decreases over infancy

Almost all bacteria have a heat shock response. Define heat shock and speculate about the temperature that might cause heat shock for a microbe that grows optimally at 10C, a microbe that grows optimally at 30C, or a microbe that grows optimally at 60C.

Heat shock response is the protection against excessive heat in order for proteins to survive as proteins can become misfolded and denatured, with sigmaH increasing affinity of RNAP to heat response regulon, which produces DnaK that is a chaperone that refolds proteins A microbe that grows optimally at 10°C: This microbe might experience heat shock at temperatures above 25-30°C, since temperatures above this range could lead to denaturation of its proteins and disruption of its cellular processes. A microbe that grows optimally at 30°C: This microbe might experience heat shock at temperatures above 40-45°C, since temperatures above this range could lead to denaturation of its proteins and disruption of its cellular processes. A microbe that grows optimally at 60°C: This microbe might experience heat shock at temperatures above 70-75°C, since temperatures above this range could lead to denaturation of its proteins and disruption of its cellular processes.

Lac Operon and diff amounts -High glucose -Low glucose, no lactose -Low glucose, high lactose

High glucose 1. No CAP (activator protein) binding to promoter 2. Repressor (LacI) bound 3. No transcription Low glucose, no lactose 1. CAP binding 2. Repressor bound since no lactose around to bind LacI to make it fall off promoter 3. no transcription Low glucose, high lactose 1. Cap binding 2. Repression relieved 3. transcription occurs

Speculate why most of the microbial stress responses involve regulation of many genes at once.

In order to survive and adapt to the stressor, microorganisms need to activate a variety of cellular processes and pathways, which often require the coordinated expression of many different genes. -by coordinating the expression of multiple genes, microbes can ensure that different aspects of their stress response are working in sync -it saves energy and at times of stress, microbes need to ration out energy.

What distinguishes apicomplexans? Define apical complex, rhoptry, microneme. Are these secretion organelles similar to bacterial secretion systems?

Is a type of parasitic protozoan with an apical complex located at one end of the cell and has secretory organelles that allow for proteins to be secreted by vesicle merging with membrane -apical complex: a specialized structure located at the anterior end of the cell. The apical complex contains various organelles, including rhoptries and micronemes, that are involved in host cell invasion and pathogenesis. -Rhoptries are secretory organelles that contain proteins that are released and take control of host cells -Micronemes are secretory organelles that bind to host cell and invade the host cell by penetrating its cell membrane There are some similarities as the secretion systems are generally used to transport proteins across the bacterial membrane or cell wall, while the apical complex and associated organelles of apicomplexans are involved in the invasion of host cells and manipulation of host cellular processes.

What is the defining feature of the kinetoplastids? What is contained in the kinetoplast?

It uses a flagella to move around and is a network of DNA inside of a large mitochondrion. The DNA is bound together in circles because it lacks the enzyme gyrase to cut them

sigmaH at high vs low temps

Low temp -small amounts of sigmaH made -mRNA folded over onto itself where SD is, blocking ribosomes and now letting them bind to make more sigmaH -heat response off High temp -melts folded mRNA and now SD region can be bound to and sigmaH made -RNAP activated so response gene is on

Do prokaryotes more often control the amount of proteins produced or protein activity? Why? (what are 4 reasons for this)

More often control the amounts of protein b/c: -substrates are limited -gene expression is set up to be controlled -mRNA have short half lives -operon organization allows coordinate regulation.

Can viruses sense and respond to their environment? Is it possible for viruses to have two-component systems? Explain how this could work or why it would not work.

No, because their spike proteins which bind to the host cell's receptors cannot sense the environment Not possible for viruses to have two-component system because two-component systems are based on a specific set of proteins that are not present in viruses, and they rely on the ability of cells to sense their external environment and generate an internal response.

We cannot put viruses on the same phylogenetic tree as bacteria, archaea and eukaryotes. Can we put all viruses on a separate, but single viral phylogenetic tree? If yes, explain how. If no, explain why not.

No, it is not possible to construct a single phylogenetic tree that includes all viruses. This is because viruses are not considered to be living organisms and are derived from the DNA of other cells. There is no common viral ancestor

4 types of parasites

Plasmodium, toxoplasma = apicomplexans Trypanosomes, leishmania = flagellates

Compare and contrast prokaryotic and eukaryotic flagella. How do their structures and function differ?

Prokaryotic and eukaryotic flagella are both whip-like structures that are used for motility -Prokaryotic flagella are outside of the cell and have a simpler, smaller structure, and made up of a protein called flagellin and consist of a single filament that rotates like a propeller -Eukaryotic flagella are made of microtubules, membrane-bound and powered by ATP -Prokaryotic flagella rotate like a propeller while eukaryotic flagella move in a whip-like motion -Eukaryotic flagella can sense the environment and participate in cellular signaling pathways while prokaryotic flagella are just for motility

What are chaperones and why are they needed in some secretion systems? Give two examples of chaperones that play a role in secretion. What do you think would happen if you deleted these factors?

Protein chaperones act to promote folding, block aggregation, disaggregate proteins, and facilitate protein degradation -they are needed in secretion systems to bind to protein and prevent it from folding during their transportation thru the cell -one example is SecB, which is a protein chaperone in the SEC system that binds to the protein to prevent it from folding while going thru Sec EYG channel -second example is SurA, a periplasmic chaperone protein that prevents protein from aggregating and recognizes and binds to the surface-exposed regions of outer membrane proteins as they are being synthesized, and then delivers the folded and assembled outer membrane proteins to the BAM complex for insertion into the outer membrane. Without these factors, proteins would randomly fold and take different shape, causing a loss of function. They can also be more vulnerable to degradation if not folded properly and might not be able to be transported across the membrane properly

Explain, from an evolutionary and geological time perspective, why there are qualities that are unique to protists, and why protists are so diverse as a group

Protozoa are dangerous microbes that have been around for a long time before multicellular eukaryotes evolved, so there has been a long time for protists to evolve and have better resistance to any attacks -Over time, protists diversified and adapted to various ecological niches, allowing them to live in almost every environment -developed a variety of adaptations that allowed them to survive and thrive in different environments, including symbiosis with other organisms, complex cell structures, and motility -The diversity of protists as a group can be attributed to their ability to adapt to various environments and ecological niches. The unique qualities of protists allow them to occupy a broad range of habitats, from marine and freshwater ecosystems to soil and host organisms

helical capsid

Rod-shaped capsomeres that form a continuous helix around the nucleic acid -capsid wraps around DNA or RNA

signal peptide (signal sequence) and signal recognition particle

SS -hydrophobic -15-30 a.a long -N-terminus of peptide embedded in membrane -does not contribute to function -present at the N-terminus of the majority of newly synthesized proteins that are destined for certain location in cell SRP: -grabs signal peptide and puts onto membrane -made of protein Ffh and sRNA ffs which recognizes the signal peptide particles

Explain how sigma factors work. What does a sigma factor recognize, why does this matter? What is a housekeeping sigma factor vs alternative sigma factor. What is the benefit of using sigma factors to regulate a regulon? Are there any limitations - what would be the problem if the cell only used sigma factors as the only way to regulate expression?

Sigma factors provide promoter recognition specificity to RNA polymerase holoenzyme, contribute to DNA strand separation, and then dissociate from the core enzyme following transcription initiation. housekeeping sigma factors are always on for essential genes Alternative sigma factors - turned on periodically for specialized genes Benefits of using sigma factors to regulate a regulon - a set of genes and full pathways can be turned off or on, if the cell only used sigma factors as the only way to regulate expression there would need to be way more proteins and it would waste material

Outline the general pattern of signal transduction systems and two component systems. What is a histidine kinase? Response regulator? How do sensors normally work?

Signal transduction system: a stressor becomes present and is sensed by a sensor protein on the cell surface who sends a signal to a response regulator protein to create a cell response i.e. chemotaxis Two component systems are proteins that sense and respond to environment Histidine kinase: sensor protein that autophosphorylates after sensing stressor and sends signal into cell via phosphorylation Response regulator: is inside cell and receives signal from the histidine kinase via its phosphorylation, activating it to bind to target genes and changing their expressions (activate or repress) in order to have the cell properly respond Sensors normally work by having an extracellular sensing domain that detects changes in the environment, and a cytoplasmic signaling domain that communicates the signal to downstream components of the signaling pathway. -will autophosphorylate self when detecting stressor and then gives that P to response regulator to activate it

similarities and differences between eukaryotic microbes and bacteria

Similarities -central dogma: DNA --> RNA --> protein -metabolism: central pathways, building macromolecules, ATP, ETC -membrane function: import channels, integrity, phospholipids Differences: eukaryotes have -genome: linear, larger, has introns, nucleus, ore complex 3D structure, transcription/translation are separate, can be diploid -cell rigidity: no PDG (soft), chitin/cellulose for strength, cholesterol, cytoskeleton actin microtubules, bigger cells -secretion of proteins: vesicles, ER/golgi, membrane traffic

Compare and contrast prokaryotic vs eukaryotic cells

Similarities -central dogma: DNA --> RNA --> protein -metabolism: central pathways, building macromolecules, ATP, ETC -membrane function: import channels, integrity, phospholipids Differences: eukaryotes have -genome: linear, larger, has introns, nucleus, ore complex 3D structure, transcription/translation are separate, can be diploid -cell rigidity: no PDG (soft), chitin/cellulose for strength, cholesterol, cytoskeleton actin microtubules, bigger cells -secretion of proteins: vesicles, ER/golgi, membrane traffic **has nucleus, cytoskeleton made of microtubule, organelles for metabolism and secretion -larger cell and genome

Describe how a small RNA (sRNA) that binds to the Shine Delgarno Sequence of an mRNA might lead to reduced protein amount in a bacterial cell.

Small srna regulate gene expression by base pairing with messenger rna, blocking the SD region so ribosomes don't bind -sRNA: doesn't code for proteins (not RNA) and finds mRNAs that have been transcribed and base pairs with mRNA so no translation and stops protein from being produced

Define the characteristics of stress to a microbe. What are some examples?

Stress is a significant change in a microbe's environment that makes growth a challenge Examples: Nutrition limitation Temperature change Change in pH, pressure, salinity Soap and detergent

types of motility -swimming, swarming, twitching, gliding, sliding

Swimming: smooth straight motion w flagella Swarming: groups of bacteria push each other and move each other with slime layer secreted Twitching: running and tumbling in jerky motion of G-, attach to surface of host cells or inanimate objects Gliding: glide on surface in flagella-like motion but w/o flagella Sliding: cells pulled away from each other

Understand the different kinds of motility, and how these kinds of movement are achieved.(swimming, swarming, twitching, gliding, sliding)

Swimming: smooth straight motion w flagella Swarming: groups of rod-shaped cells push each other and move each other with slime layer secreted Twitching: running and tumbling in jerky motion of G-, attach to surface of host cells or inanimate objects Gliding: glide on solid surface in flagella-like motion but w/o flagella Sliding: cells pulled away from each other, can adjust their length (extend to attach and then retract to move)

Compare and contrast the various secretion systems. What elements are shared between secretion systems? What elements are different? Why do you think there's so many kinds of secretion systems?

The SEC (Secretion) system is the most common pathway used for protein secretion in bacteria. It is responsible for the translocation of unfolded proteins across the cytoplasmic membrane. The SEC system is composed of the SecA protein, which binds to the signal peptide of the protein being translocated, and brings it thru the SecYEG membrane channel (a protein-conducting channel through which the protein can pass) after hydrolyzing ATP with SecB binding to protein so it doesn't fold while moving thru channel -found in most bacteria and is essential The TAT system is a secretion system that is used to translocate folded proteins across the membrane and is rarely essential and not found in all species. The TAT system is composed of the TatABC membrane complex, which recognizes proteins with a specific signal sequence containing a pair of consecutive arginine residues, and TatD, a cytoplasmic chaperone that helps to maintain the stability of the transported proteins. The BAM system is responsible for the secretion of outer membrane proteins in Gram-negative bacteria going thru two membranes. The SEC complex brings in unfolded protein into periplasm where chaperones bring it thru BAM complex on outer membrane. The BAM complex consists of five proteins, BamABCDE, which form a complex that recognizes and inserts beta-barrel proteins into the outer membrane.

What is the Sec system and how are proteins targeted there? Where is the signal sequence encoded? Does it matter if its at the N-terminus or C-terminus of the gene? Explain why.

The SEC system is the secretion of unfolded proteins that is found in most bacteria and is essential -the N-terminal signal sequence (hydrophilic) is targeted and recognized by protein SecA that binds to it and brings it through the SEC E/Y/G membrane channel where SecB the protein chaperone binds to the protein to ensure it doesn't fold while moving thru membrane. -the signal sequence is encoded at the N-terminal end of the protein, where the oldest made amino acids of the protein are located -signal sequences located at the N-terminus of the protein are more efficient and specific in targeting proteins to the Sec system than those located at the C-terminus. This is because the Sec machinery recognizes the signal sequence as the protein is being synthesized, and the N-terminal location allows for more efficient recognition and targeting of the protein to the membrane

In the last few lectures, we've discussed several regulatory systems. Which is the most simple (least parts) to turn on? Which is the most complex? a. Lactose utilization b. σΗ regulon / heat shock response c. stationary phase d. quorum sensing in gram positive

The most simple regulatory system to turn on would be lactose utilization. This system involves the LacI repressor, which binds to the operator sequence of the lactose operon and prevents transcription of genes involved in lactose utilization. When lactose is present in the environment, it binds to LacI and causes a conformational change that prevents it from binding to the operator, allowing transcription to occur. This system involves only one regulatory protein (LacI) and a single operator sequence, making it relatively simple. The most complex regulatory system to turn on would likely be the σΗ regulon / heat shock response. This system involves multiple genes and regulatory proteins that work together to respond to heat stress. The system includes the heat shock sigma factor σΗ, which directs RNA polymerase to transcribe genes involved in heat shock response, as well as chaperones and proteases that help protect proteins from denaturation and promote refolding. This system also involves multiple regulatory pathways, including the two-component system and alternative sigma factors, which help to coordinate the response to heat stress. Overall, the σΗ regulon / heat shock response involves multiple proteins, pathways, and feedback loops, making it a relatively complex regulatory system to turn on.

Scientists can alter the DNA in a promoter so that a transcriptional activator becomes a transcriptional repressor. Explain how this could be done.

The sequence that the activator normally binds to is now placed in front of the promoter, so now when it is in its active form and binds DNA, it is physically blocking the transcription machinery and represses that operon.

specialized systems

Type I: ABC exporter, transporter takes protein out and this is for proteins lacking signal sequence Type II: like BAM, two-step secretion with protein moving into periplasm and then Om Type III: contact-dependent secretion with the injection of genetic material like a virus Type IV: conjugal transfer system with 2 bacteria connected via pilus Type V: autotransport with protein going thru 2 membranes but without help of proteins Type VI: contact-dependent secretion with injection of proteins into target cells

mutualism parasitism Commensalism

bacteria benefits and host benefits A relationship between two organisms of different species where one benefits and the other is harmed A relationship between two organisms in which one organism benefits and the other is unaffected

What are viruses made of? Can they have all 4 macromolecules (nucleic acid, protein, lipid, carbohydrate) in the virion? How can you classify viruses based on the components of the virion alone (i.e. without knowing any sequence information, or life cycle details).

Viruses are generally composed of protein shell capsid that encloses genetic info (DNA or RNA) with spike proteins facing outside that bind to host cell's receptors -can also have an envelope made of lipid bilayer membrane that is made out of host's lipids Viruses can have all 4 macromolecules, depending on the host they infect -when there is no membrane or helical capid, the structure has proteins and nucleic acids -when there is a membrane but not helical capsid, there are nucleic acids and an envelope (lipid) - when there is no membrane but a helical capsid, there is a nucleic acid and protein -when there is a membrane and helical capsid, there are nucleic acids and envelope (lipid)

When an E. coli culture shifts between glucose and lactose utilization, a lag phase is observed. However, when E. coli is shifted from 30°C to 40°C, the growth rate continues swiftly but no lag is observed. Using your knowledge about how these systems are regulated, explain why.

When E. coli is shifted from glucose to lactose utilization, there is a lag phase before the cells can begin to grow and divide on lactose. This is because E. coli needs to activate the genes required for lactose utilization, which involves de-repression of the lactose operon and activation of the necessary transporters and enzymes. This process takes time and requires the synthesis of new proteins, which contributes to the lag phase. In contrast, when E. coli is shifted from 30°C to 40°C, there is no lag phase observed. This is because the heat shock response is a pre-existing stress response system that is already activated at low levels even at the optimal growth temperature of 30°C. When the temperature is increased to 40°C, the heat shock response is rapidly upregulated to protect the cells from protein denaturation and other stresses associated with high temperature. This response involves the activation of chaperones and proteases, which are already present in the cells and can be rapidly synthesized to provide protection against the stress.

cell stress

a change in conditions that reduces growth or survival -proteins sense stressor and produce cellular signal that reprogram specific genes to respond with a feedback mechanism inactivating signal and response when stressor no longer present -conditions that microbes find stressful: temperature, pH, pressure, salinity, low nutrients, O2 toxicity, competition, soap/detergent (membrane stress)

What is the advantage of allosteric regulation? How can negative feedback regulate a biosynthetic pathway?

advantage of allosteric regulation allows the control of specific enzyme actives, can change based of of environment. negative feedback inhibition occurs when the products of a biological reaction stops the reaction from continuing to occur when there is no stressor detected, allowing cell not to waste any energy

Agglutination

clumping of red blood cells after a virus bridges 2 red blood cells -virus clumps multiple RBC -spin blood: ---no virus = no clumping of cells = red sediment ---virus= clumps of RBC means virus is bound = hazy red liquid

What is a hemagglutination assay and how does it work? Do you think a similar assay could / would be used to quantify bacteria? Why or why not?

clumping of red blood cells after a virus bridges 2 red blood cells by binding to its receptors -works by spinning the blood and if the red blood cells clump, that means that a virus is attached to them while if there is no clumping and a sediment forms, there is no virus It is unlikely that a similar assay could be used to quantify bacteria, as bacteria do not typically bind to or agglutinate red blood cells since they do not have a spike protein that recognizes a cell's receptors

chemotaxis

directed cell movement in response to chemicals which can be either attractants or repellants -attraction of bacteria to chemicals as cell's movement directed by controlling frequency of runs and tumbles (increase frequency of tumbling when in wrong direction)

spores

dormant form of cells that can survive harsh conditions

virions

individual virus particles -all components of virus made by host cell -capsid protein -genome has RNA -spike protein that can't sense environment and binds to host cell's receptors

prions

infectious protein particles that do not have a genome -not a virus -wrongly contagious folding that binds to normal folded proteins, making those also wrongly folded, thus disrupting function

heat shock

protection against excessive heat as proteins can become misfolded and denatured 1. sensor: sigmaH increases affinity of RNAP for ptomoters of heat shock regulon 2. signal: is denatured proteins - product of heat shock regulon is DnaK, a protein chaperone that facilitates proper protein folding, binds to and target sigmaH for degradation --> at high temps, denatured proteins accumulate and bind DnaK to divert it from sigmaH 3. response genes: heat shock regulon has 250+ genes activated by sigmaH with some encoding proteins that change cell membrane composition, protein chaperone DnaK refold denatured proteins, proteases degraded damaged proteins 4. feedback control: when no more unfolded proteins around, cell decreases production of heat shock proteins w DnaK proteins now free to bind to sigmaH to protease FTSH for degradation and turns system off

Compare and contrast the strategies used by transcriptional activators and repressors. What aspects are the same? What aspects are different?

repressors: -located between the -10 and the -35 -block transcription that would normally happen if it wasn't present activators: -usually located before the -35 and -10 -increase the transcription from a promoter -promoter is probably weak -the -10 and -35 are not very similar to the ideal/consensus -sigma can't recognize the promoter, so they can't initiate transcription there -activators act either by promoting the binding of RNA polymerase, or by helping it to start transcribing better Similarities: -Both bind to specific DNA sequences called DNA control regions -Both can recruit other proteins that either enhance or inhibit gene expression. -Both can interact with the RNA polymerase to influence the initiation of transcription

TAT system -3 steps

secretion of folded proteins -not found in all species, rarely essential 1. membrane protein channels TATB/TATC recognize signal peptide and it enters periplasm from cytoplasm 2. TATA activated by ETC and PMF (needs E) is a multimer on membrane and protein enters it 3. SPase1 allows for protein to be released out into periplasm with signal peptidase chewing up additional subunits and whatever protein was attached to

BAM system -3 steps

secretion of outermembrane proteins (G-) where protein can go thru 2 membranes 1. proteins going to OM carry N-terminus signal sequences that is recognized and cleaved off by Sec appartus via SEC system with unfolded protein passing thru channel into periplasm ---unfolded protein passes inner membrane via SEC 2. Nascent proteins bound by chaperone protein Skp to prevent them from folding in periplasm and the chaperone escorts protein to dedicated protein complex of OM called BAM 3. BAM inserts cargo into outermembrane by folding protein with chaperone help and inserting protein into outermembrane DegP protease degrades misfolded proteins or damaged proteins in order to prevent entrance to OM

SEC system -5 steps

secretion of unfolded peptide -exports most periplasmic and outermembrane proteins of G- and secreted proteins of G+ -found in most bacteria, essential (most secreted proteins) 1. N-terminal signal sequence (hydrophilic) is recognized by SecA protein, which binds to it 2. SecA brings SS where chaperone SecB binds to protein and prevents protein from folding when going thru channel 3. SecA brings SS to cell membrane, translocating it, where it goes thru proteins Sec E/Y/G in membrane, and this allows hydrophilic and hydrophobic proteins to thread their way thru hydrophilic inner surface of channel 4. SecA has to hydrolyze ATP to allow 20 a.a thru the channel 5. Signal peptidase clips off protein's SS as it passes thru Sec E/Y/G channel via enzymatic proteolysis with the protein now folded and can function outside of cell -passage thru channel needs PMF or ATP

histidine kinase

sensor protein that phosphorylates its own histidine residue after sensing stressor

liposome

small membrane bounded transport vesicles -exist on own, drug delivery, miscelle shaped -injected gene inside and transported to target -covid vaccine

Bacteria express different things during log phase and stationary phase. What kinds of genes would you expect to be expressed a) mostly during log phase, b) mostly during stationary phase c) similarly during both? Why?

the gene expression profile of bacteria changes dramatically during the transition from log phase to stationary phase, reflecting the shift in priorities from growth to survival During log phase, bacteria are actively dividing and replicating their DNA, requiring a significant amount of energy and resources to support growth. As a result, during log phase, we would expect bacteria to express genes involved in DNA replication, cell division, and metabolic pathways that generate energy and building blocks for cellular growth During stationary phase, the rate of bacterial growth slows down as nutrients become depleted and waste products accumulate, leading to a shift in gene expression towards survival mechanisms such as stress response and nutrient scavenging. We would expect bacteria in stationary phase to express genes involved in stress response pathways, such as chaperones, proteases, and DNA repair enzymes. Additionally, genes involved in nutrient acquisition and storage There are also some genes that may be expressed similarly during both log and stationary phases, such as those involved in basic cellular processes like transcription and translation, as well as genes involved in regulation of gene expression, such as sigma factors and global regulators. Additionally, some stress response genes may be expressed in both log and stationary phases to provide a baseline level of protection against stressors, even when bacteria are actively growing.

regulon

the set of genes controlled by a given sigma -same sigma factor gives genes that have similar sequences at the -35 and -10 regions

How do microbes respond to stress? Outline the 4 general steps involved in any stress response.

transmembrane proteins sense signal and transmits chemical signal to inside the cell to turn on certain genes 1. Sensing stress: can be absolute or dynamic 2. Signal transduction: chain reaction with protein modification 3. Response: gene regulation increase or decreased 4. Resolution: feedback inhibition with turned on genes turned off

Quantifying viral replication PFU

virions are too small to scatter light, to see by microscopy, to see a colony plaque-forming units: concentrations of viral suspensions measured by the number of plaques -host cells (bacteria or eukaryote) on plate, add viral particle, dead cells are the white spots, plaques -benefit: counts infectious virions -downside: slow method