MICROBIOLOGY EXAM 1

What are endospores? Describe sporogenesis?

A bacterial endospore is a complex, dormant structure formed by some gram-positive bacteria (like anthrax). They occur in various locations and are resistant to environmental conditions, such as heat, radiation, chemicals, and desiccation (drying out). Spores have a thin covering called exosporium, which is a thick layer of protein coats. Beneath those coats is the cortex, or thick peptidoglycan with a nucleoid and ribosomal core. A spore is so resistant because it contains calcium with dopicolinic acid, it's small, acid soluble, and has DNA binding proteins (SASPS), and has a dehydrated core. Once the spore has been germinated it enters vegetative form, which is where it becomes disease causing. Germination occurs in treatments like heating. -Sporogenesis is the production of spores. The process goes as such 1. Axial filament formation 2. Septum formation and forespore development 3. Engulfment of forespore 4. Cortex formation 5. Coat synthesis 6. Completion of coat synthesis, increase in refractility and heat resistance 7. Lysis of sporangium, spore liberation

How would one maintain a continuous culture in the laboratory? Why would someone want to maintain a continuous culture?

A continuous culture allows for microbial growth in the exponential (log) phase for an extended period of time at a constant biomass concentration. This type of culture maintains conditions for growth by continual provision of nutrients and removal of wastes. A continuous culture uses a chemostat or turbidostat. In the chemostat an essential nutrient is in limiting quantities and the dilution rate is the rate at which the medium flows through the vessel relative to the size of the vessel. Also, the rate of incoming medium is equal to the removal of medium from the vessel. The dilution rate of the turbidostat varies because the flow rate of media through the vessel maintains a predetermined turbidity or cell density. The turbidostat is most stable and effective at high dilution rates, and none of the nutrients are limiting.

What is the structure of the outer membrane? What is its function? What are porins? Why is lipid A called an endotoxin?

A. The Outer Membrane Structure: Only gram-negative bacteria have an outer membrane(OM) and it lies outside of the thin peptidoglycan layer (Image below). It's linked to the cell by Braun's lipoprotein (most abundant in OM, joined to the peptidoglycan covalently and is embedded in the OM by its hydrophobic ends. There are large complex molecules on the OM (lipopolysaccharides: LPS) which have both lipid and carbohydrate, consisting of three parts: Lipid A Contains two glucosamine sugar derivatives ea. with three fatty acids phosphate or pyrophosphate attached. The fatty acids are embedded in the OM while the rest sticks out like some sort of peach fuzz. The core polysaccharide Joined to Lipid A and made of 10 sugars (many unusual) The O-chain or O-antigen Polysaccharide chain extending from the core. Aslo has unusual sugars and varies in composition between bacterial cells. B. What is its function? Importance of LPS contributes to negative charge on cell surface helps stabilize outer membrane structure may contribute to attachment to surfaces and biofilm formation creates a permeability barrier The geometry of LPS and interactions other LPS beside it may restrict bile salts, antibiotics, and other toxic substances. protection from host defenses (O antigen) O-antigen elicits an immune response from the infected host and the response is antibodies that bind to strain-specific forms of LPS that caused the response. Many bacteria can rapidly change their O-chain to thwart host defense. Why is Lipid A an endotoxin? can act as an endotoxin (lipid A) Causes symptoms that can arise in infections by Gram-neg pathogens. If Lipid A gets into the bloodstream, septic shock develops, which there isn't direct treatment for.

Mycoplasma genitalium, a cell wall-less bacterium that grows in human serum, lacks the ability to synthesize fatty acids. How do you think M. genitalium makes its cell membrane?

Although it is not known why species like M. Genitalium cannot lack the ability to synthesize fatty acids, it is thought that it can still make a cell membrane due to its ability to synthesize membrane phospholipids and glycolipids from their own hosts. This results in an overflow of cholesterol in to its host as a buffer of membrane fluidity.

How can non-consecutive amino acids form the active site of an enzyme? What is the function of disulfide bonds in proteins?

Although the proteins are not consecutive, when folded into the tertiary structure, the amino acids fold together to create an active site for the enzyme.

If metabolic functions require a relatively narrow pH, how can bacteria exist in environments with a wide range of pH values?

Bacteria exist in environments with a wide range of pH values because they have mechanisms to maintain an internal pH value near neutrality. For example, when there is a minimal change to external pH some bacteria use an antiport transport system that exchanges potassium for protons. When the external pH is too acidic bacteria can utilize the acidic tolerance response which allows the ATPase enzyme to pump protons out of the cell at the expense of ATP. The acidic tolerance response also contributes to the synthesis of proteins such as acid shock proteins and heat shock proteins that prevent the denaturing of proteins and aid in refolding denatured proteins.

Describe how bacterial fermentation causes milk to sour.

Bacterial fermentation causes milk to go from a relaxing drink to sour and foul-tasting. The milk is initially inhabited by bacteria that create acidic conditions, mainly through lactic acid fermentation. As the milk becomes more acidic, these bacteria are killed by the acidity and replaced by better-adapted bacteria that make the milk even more acidic. This acidity makes the milk sour and causes it to coagulate. As the milk still becomes more acidic, even these bacteria die and yeasts and molds become more dominant in the environment and degrade this lactic acid. Then protein-digesting bacteria become more prevalent and, through putrefaction, degrade proteins into amine compounds such as cadaverine that give it an even more terrible odor and bitter flavor. Bacteria in milk such as lactobacilli break down lactose by fermentation, producing lactic acid. Acidic substances have a sour taste. The acidic conditions created by lactic acid make the milk protein, casein, turn to curd, and this further notifies us to not drink it.

What is the difference between growth and reproduction?

Cell growth is the process of a cell increasing its cytoplasmic and organelle volume (G1 phase), as well as increasing its genetic material (G2 phase) following the replication during S phase. Cell reproduction is the actual division of the cell that produces daughter cells that will develop and continue the cycle.

A microbiologist describes an organism as a chemoheterotrophi, aerotolerant, mesophilic, facultatively halophilic bacillus. Describe the organism's metabolic and structural features in plain English.

Chemoheterotrophic means that it is an organism that derives its energy by ingesting intermediates or building blocks that it is incapable of creating on its own. Supplement. Chemotrophs are organisms that obtain energy through chemical process called chemosynthesis rather than by photosynthesis. Mesophilic means it must be an organism that grows optimally between 20-45 degrees C. Bacillus means it rod-shaped. Halophilic means it can survive in high salt concentrations. Aerotolerant means it can grow with or without oxygen but since it facultative it will grow better in the presence of oxygen.

A new chemotherapeutic drug kills bacteria, but not humans. Discuss the possible ways the drug may act selectively on bacterial cells.

Chemotherapeutic drugs can successfully kill bacterial cells but spare human cells, through taking advantage of some of the distinctive differences between these two cell types. The two main ways include the drug's effects on either the bacterial cell wall or the metabolic pathways of the bacterial cell. If the chemotherapeutic drug is to effect a cell wall (directly affecting and inhibiting peptidoglycan synthesis in cell walls); the reason this would lyse the bacteria is due to the fact that human cells do not have this cell structure and therefore it would not be affected. By destroying the cell walls the bacteria can lose its shape and be destroyed by osmotic pressure. Bacterial metabolic pathways also differ from that of human cells (specifically eukaryotic). Sulfa drugs in this case can inhibit the intake of nutrients necessary for purine synthesis.

European manufacturers have included chlorhexidine in their toothpastes and mouthwashes for many years, but there has been resistance to this practice in the US. Would you favor or oppose such a move? Why?

Chlorhexidine, is a topical antiseptic that is used for skin disinfection before surgery and to sterilize surgical instruments. Chlorhexidine when used in the oral route (mouthwash or toothpaste) is used to treat gingivitis (only serious cases though - requires prescription in US). Gingivitis is caused by the bacteria that grow in the coating (plaque) that forms on teeth between tooth brushings and can lead to oral decay. Chlorhexidine destroys and lysis the bacteria, thereby preventing the gingivitis from occurring. It does serve its purpose well, especially in a mouthwash form but toothpaste is highly advised against because the remnants can remain in one's mouth if not thoroughly washed out. This product should only be prescribed in serious oral infection cases, because chlorhexidine is a powerful agent that can go too far especially if it is not rinsed out, it is not recommended for any treatment on children, and it can cause further oral problems such as destruction of tooth fillings (metal and acrylic cavity fillings or sealants) and can cause serious periodontitis, which is a common but serious gum infection that damages the soft tissue and destroys the bone that supports your teeth; this can cause teeth to loosen or lead to tooth loss.

What are cofactors and coenzymes? How do they participate in the chemical reactions in a cell?

Cofactors are the nonprotein component of an enzyme. They include metal ions as well as a variety of organic molecules. When a cofactor is combines with the apoenzyme, it is called a holoenzyme. A coenzyme is a type of cofactor that is attached loosely to the apoenzyme. This way it can dissociate from the apoenzyme after products have been formed, and be used again. Many of our vitamins react as coenzymes. Coenzymes often will carry a product of the reaction to another enzyme or transfer chemical groups from one substrate to another.

If you wanted to know how many dead cells were in a culture, what would you do?

Direct counts on membrane filters can tell you how many cells are presev bgnt, and with certain dyes distinguish living from dead cells. Viable counting focuses more on how many cells are alive in a culture.

What is end-product inhibition? Explain allosteric activation.

End-product inhibition is a type of feedback inhibition that regulates metabolism taking place in the cell where the end product of a pathway will, when manufactured to a certain amount, react with the pacemaker enzyme (regulatory enzyme) to stop the pathway from producing more of the end product. Allosteric activation is another type of regulation of metabolism in the cell where allosteric enzyme (regulatory enzyme) is altered by the allosteric effector through binding reversibly by noncovalent forces to a regulatory site (separate from catalytic site) and causes a shape change to the catalytic site of enzyme changing the affinity of that the enzyme has for the substrate (positive - increased reacting; negative - decreased reacting)

It is generally estimated that in a typical cell 1000-2000 different chemical reactions are catalyzed by enzymes. Why do you suppose it is necessary that each of those reactions have its own special enzyme?

Enzymes are proteinaceous substances that makes life possible. They are needed for every chemical reaction that occurs inside the human body. Enzymes are specific in what they functions and on which substrate and at what pH level they work. Each enzyme requires a particular temperature in order to functions optimally. Thus, each reaction requires a special enzyme for their activity and to carry out the body metabolism in a right way.

Explain how enzymatic reactions are coupled to supply useful energy.

Enzymes have the ability to excite or inhibit reactions in metabolic processes. They also control the speed of the enzymatic activity by increasing the concentrations of substrates at active site of enzyme. Enzyme Inhibitors: Competitive Inhibitor: attaches to the active site. Has a similar structure to substrate and compete to inhibit reaction Noncompetitive Inhibitor: binds to enzyme at other sites. Changes the shape of the enzyme to slow activity Regulation: -Allosteric effector: bind to to regulatory state to change shape on enzyme so that the proper substrate can bind to catalytic site (positive= increases activity; negative= decreases activity).

Enzymes can catalyze chemical reactions. How do enzymes increase the rate of a reaction? What constitutes the active site? For an enzyme-catalyzed reaction, what are the Km and Vmax?

Enzymes speed up the rate at which a reaction proceeds toward it's final equilibrium by lowering the activation energy increasing concentrations of substrates and the active site of the enzyme. They increase by orienting substrates properly with respect to each other in order to form the transition-state complex. And the induced fit model for enzyme-substrate interaction. The active site is the region of an enzyme where substrate molecules bind and undergoes a chemical reaction. The active site must be able to form temporary bonds with the substrate and catalyze its reactions.The rate of reaction when the enzyme is saturated with substrate is the maximum rate of reaction, Vmax. The relationship between rate of reaction and concentration of substrate depends on the affinity of the enzyme for its substrate. This is expressed as the Km of the enzyme, an inverse measure of affinity.

Members of the pathogenic bacterial genus Haemophilus require NAD+ and heme from their environment. For what purpose does Haemophilus use these growth factors?

Growth Factors: NAD+ and heme; NAD+ serves as an oxidizing agent and is an electron carrier. Heme houses iron and is essential for energy generation in the electron transport chain (electron shuttle) and detoxification of host immune effectors. It is the factor that allows a bacteria to cause disease. Both of these factors are essential for creating energy. As far as their influence for growth, heme allows for the bacteria to survive in a host and supports essential enzymes.

Why do changes in H+ concentration affect cell growth?

Hydrogen ions play an important role in the cell growth: The changes in hydrogen ion concentration are intimately tied to the charge of proteins side chains. This charge state, in turn, affects the activity of enzymes as well as their folding and even in localization. Further, the famed Adenosine Tri-Phosphate (ATP) synthases that churn out the ATPs production, which power many cellular processes that are driven by gradients in hydrogen ions across membranes.

What is hydrostatic pressure? Why can microorganisms withstand high hydrostatic pressures? How is hydrostatic pressure harmful to microorganisms?

Hydrostatic pressure is the pressure that is exerted by a fluid at equilibrium at a given point within the fluid, due to the force of gravity, and it increases in proportion to depth measured from the surface. The reason for this is the increasing weight of fluid exerting downward force from above. Hydrostatic pressure is known to cause morphological, biochemical, and genetic alterations in vegetative microorganisms, generally because most microorganisms have adapted to 1 atm of pressure. Generally, yeasts and molds are most sensitive to hydrostatic pressure, while gram-positive microorganisms are most resistant, possibly due to their cell wall structure, and gram-negative microorganisms are moderately sensitive. Bacterial spores are extremely resistant to hydrostatic pressure. There are also Barophiles, which are adapted to and even require the high pressures (hydrostatic pressure for the ones on the ocean floor).

*When the local drinking water is believed to be contaminated, area residents are advised to boil their water before drinking. Often, however, they are not told how long to boil it. As a student of microbiology, what might be your recommendation?

I would recommend that the residents intermittently boil their water for 30 minutes and then cool it so they can be protected against potential endospores contaminating the water. The cooling of the water would allow a period of time for some of the endospores to germinate into vegetative cells that would then be destroyed in the next round of boiling. By repeating this sequence a few times, more and more endospores are destroyed and the water would eventually be sterilized.

If ATP is such an important energy source in bacteria, why do you think it is not added routinely to the growth medium for these organisms?

If ATP is such an important energy source in bacteria, why do you think it is not added routinely to growth media? The ATP-generating reactions of glycolysis, the citric acid cycle, and the redox reactions of the electron transport chain have other important functions as well. Many of the intermediates of the pathways listed are precursors for anabolism (the synthesis of larger, more complex molecules from smaller, simpler ones). For instance oxaloacetate is the precursor to many amino acids. The proton motive force generated by the ETC is not only used to synthesize ATP but it is also very important for providing the energy needed for secondary active transport. Thus, by simply supplying the cells with sufficient ATP, these other important functions of ATP-generating reactions would not be carried out. In addition, I know of ATP at high enough concentration acting as a regulatory molecule in pathways like the citric acid cycle. While supplying cells with ATP may provide them with energy needed for some important functions, such as active transport, it may at the same time inhibit other important pathways.

If an organism can live in an anaerobic environment, does that mean that the organism uses oxygen to grow? Why or why not?

If an organism lives in an anaerobic environment it does not have to have oxygen to grow. In fact, it could possibly be different types of microorganisms. A facultative anaerobe does not require oxygen to grow, however it grows better in its presence. An aerotolerant anaerobe grows equally well in presence or absence of oxygen. Therefore, overall the organism can use oxygen to grow yet it does not require it if it can live in an environment lacking oxygen.

Faculative anaerobes can live under either aerobic or anaerobic conditions. What metabolic pathways allow these organisms to continue to harvest energy from sugar molecules in the absence of oxygen?

If oxygen is not available, facultative anaerobes are able to use anaerobic pathways to produce ATP, but not as much ATP is produced

Is it correct to say that energy is transferred from one molecule to another by oxidation-reduction reactions? Why or why not?

It is correct to say that electrons are transferred from one molecule to another by oxidation-reduction reactions. Energy from the redox reactions can be stored and later used for ATP synthesis but energy is not being transferred from molecule to molecule. For example, the electron transport chain is a series of sequential redox reactions carried out by electron carriers. Each carrier in the chain is reduced allowing the electrons to be transferred to the next carrier and then reoxidized to accept more electrons. The first carrier in the chain has the most negative standard redox potential making it a better electron donor. The terminal electron acceptor has least negative standard redox potential making it a better electron acceptor. The electrons are transferred from carrier to carrier via redox reactions, releasing energy along the way, until the terminal electron acceptor is reached. This is followed by the formation of ATP. (Carriers can be NAD, NADP, FMN, FAD, Cytochromes, coenzyme Q, etc.)

Why is the reducing power found in NADH and FADH2 considered a form of energy?

NADH and FADH2 help form the PMF (proton motive force), which is then utilized by ATP synthase to start ATP synthesis. Additionally, they have a negative E'0 and are able to be used to conserve energy. They also serve as a form of energy because of their ability to donate H+, which would make another molecule more rich in energy, thus transferring energy.

A medical microbiological lab report indicates that a sample contained a biofilm, and that one species in the biofilm was identified as Neisseria gonorrhoeae. Is this strain of Neisseria likely to be pathogenic? Why or why not?

Neisseria gonorrhoeae is an obligate human pathogen, that is identified as being a gram-negative, oxidase-positive, diplococcus strain (they form diplococci rods), and has been cited as a causative agent of gonorrhea (an STD). This particular species is always considered to be pathogenic, as it is not considered normal flora under any circumstance. Similar to other bacterial species of this nature (Neisseria), it does hold the potential to create a bacterial biofilm. These biofilms are thin, slimy, biological communities closely interrelated in a matrix, and are capable of colonizing on other tissues and even inorganic surfaces (such as certain metals), and have many advantages compared to free moving 'unattached' bacteria. One of these advantages is increased resistance to antimicrobial agents. In addition to that, the individual cells of this species of can alter their surface structure as well as its pili and protein antigens, allowing this organism to have the ability to evade the host immune response; with the aid of a biofilm that can attach to epithelial cells this organism is capable of infecting a wide variety of mucus secreting areas within the body (primarily uro-genital organs).

How do organisms determine which of their catabolic pathways to use? How does catabolism depend on environmental factors?

Organisms choose which catabolic pathway to use based on the materials had by said organism. Due to the fact that energy through ATP is scarce and used only when needed, it would be inefficient of the organism to use a pathway that requires materials that the organism does not have. For example, the Entner- Doudoroff Pathway requires the intermediate KDPG. In contrast, the Pentose Phosphate Pathway requires pentose sugars as well as glucose-6-phosphate. Lastly, the Embden-Meyerhof Pathway needs phosphorylate glucose in order begin the 6-carbon phase. Catabolism is also determined partly by environmental factors. This is due to the fact that certain organisms undergo specific pathways based on the presence and absence of oxygen. The TCA cycle occurs in aerobic respiration for aerobic bacteria, free-living protists, and fungi. In contrast to the TCA cycle, the Embden-Meyer Pathway can occur with or without the presence of oxygen for all major organisms. The presence or absence of oxygen plays a huge role in determining which pathway will be used.

A bacterium has been isolated from a patient and identified as a gram-negative rod. Knowing that it is a human pathogen, what structures might it have? Explain your reasons for each choice.

Outer membrane-Inner membrane-periplasmic space (large)- thin peptidoglycan layer

Explain the following statement: The structure of all biomembranes depends on the chemical properties of phospholipids, whereas the function of each specific bio-membrane depends on the specific proteins associated with that membrane.

Phospholipids consist of a hydrophilic (or 'water loving') head and a hydrophobic (or 'water fearing') tail. Phospholipids like to line up and arrange themselves into two parallel layers, called a phospholipid bilayer. This layer makes up your cell membranes and is critical to a cell's ability to function. All cells contain this layer to control what goes in and out of the cell. It is crucial to all cells and the structure is consistent in various classes of organisms. However, each organism has properties or functions that are unique to itself and that all organisms do not do. This requires various specific proteins to be present to carry out the functions. For example, an microbe that is able to survive in extremely salty locations will have different proteins that keep their salt concentration inside the cell from getting too high.

*Why is phosphorylation an important part of metabolism?

Phosphorylation is an important aspect in metabolism because it is a means of post-transcriptional regulation that affects the regulatory enzyme quickly and thus is a rapid means of altering the flux through the pathway containing that enzyme. Phosphorylation can either turn "on" or turn "off" a particular enzyme, and it can sometimes even contribute to partially enhancing or diminishing the activity of an enzyme depending on the level of phosphorylation of a multi-subunit enzyme.

Name the significant physical factors in a microorganisms non-nutrient environment. Name the significant chemical factors.

Physical factors: temperature, pH, water activity, pressure, radiation Chemical factors: available oxygen, carbon, and other organic or inorganic molecules for energy and carbon sources Ch.7 and 11 Extremophiles: microbes that grow in extreme environments Osmophiles: microbes adapted to extreme hypertonic environments Osmotolerant: can grow at a variety of water activity, but optimal at higher levels Halophiles: extreme saltiness Acidophiles: like acidity, low pH 0-5.5 Neutrophiles: neutral pH 5.5-8 Alkaliphiles: more basic, pH 8.0-11.5 Psychrophiles: low temps 0-15℃ Psychotrophs: 0-35℃ cause for food spoilage Mesophiles: 15-45℃ Thermophiles: 45-65℃ Hyperthermophiles: high temps 55-113℃ Barotolerant: low pressures, but not too low Plezophilic (barophilic): higher pressures, higher than 1 atm for maximum growth Photolithoautotrophs (Photoautotrophs): light, CO2, inorganic molecules Chemolithoautotrophs: inorganic molecules, CO2 Chemoorganoheterotrophs (Chemoheterotrophs): organic molecules Photoorganoheterotrophs: light and organic molecules Chemolithoheterotrophs: inorganic for energy and organic for carbon

Describe two ways in which energy is used to move ions and solutes against a concentration gradient.

Primary active transport: uses hydrolysis of ATP to power transport; they are uniporters (move a single molecule across the membrane). This energy uses ATP-binding cassette transporters (ABC) to use for import and export of substances. Secondary Active transport: uses potential energy of ion gradients across a membrane to power active transport; cotransporters (both same direction = symport / different direction = antiport). This transport system includes MFS proteins (major facilitator superfamily) to use ion gradients.

What are five major differences between prokaryotic and eukaryotic cells?

Prokaryotic cells lack a membrane-bound nucleus. Prokaryotic cells do not have a cytoskeleton, membrane bound organelles, and internal membranous structures like the endoplasmic reticulum and the Golgi apparatus. Prokaryotic cells have a cell wall, in bacteria cells the cell wall is composed of peptidoglycan, but in archaea cells peptidoglycan is absent from the cell wall. Eukaryotic cells are generally larger in size and more complex compared to prokaryotic cells. The structure of DNA in eukaryotic cells is open, linear, and double stranded. The structure of DNA in bacteria cells is closed, circular, and double-stranded.

A hand-sanitizing solution inoculated with 2 X 10^6 organisms of Staphyloccous aureus exhibits a D-value of 5 minutes. What will the D-value be is 10 times more S. aureus is added?

The D-value or the decimal reduction time is the amount of time required to kill 90% or 1 log of the microbial population. It is independent of the inoculum size. The D-value will be 5 minutes even if 10 times more S.aureus is inoculated.

If two cultures of E coli were growing in media of the same composition, one exposed to air and one anaerobically, which would have the larger biomass when growth ceased? Why?

The culture left growing in the air would have a larger mass because 1. Most microorgs are aerobic, and they could have a higher growth rate due to the presence of oxygen being able to produce more energy and 2. They have more exposure to microorgs in the air, and so new colonies may begin to grow, increasing the size of the overall biomass.

Subtilisin is a bacterial protease that can cleave (break) any peptide bond, regardless of the specific amnio acids involved. Trypsin, on the other hand, splits peptide bonds only on the carboxyl side of lysine and arginine groups. What differences might you expect in the active sites these two enzymes?

The difference between active sites on enzymes that can break any peptide bond vs. those that break only a specific type of bond is the number of active sites per particle. On average, enzymes like Trypsin have one active site per particle. On the other hand, proteases are specific for certain amino acids and can hydrolyze the amino acids on the carboxy or amino side of the peptide bond. These enzymes may have more than one active site, which coordinate the hydrolysis of the peptide bond.

Why are most antimicrobial chemical agents disinfectants rather than sterilants? Why might it be necessary to employ procedures like use of dilution and in-use tests?

The function is to create cystine which connects the polypeptide chains, promotes stability, and determine structure. They are disinfectants rather than sterilants due to the fact that it is extremely difficult, time consuming and expensive to sterilize on a regular basis in a home. It is also much more harmful to objects and humans to sterilize. It is important because it allows a more accurate determination of proper disinfectant concentration for a particular situation. One that is diluted so much that it kills most bacteria.

*Isolation of some membrane proteins requires the use of detergents; isolation of others can be accomplished with high salt solutions. What types of membrane proteins require detergents? What types are isolated with high salt solutions? Why?

The isolation of integral/transmembrane proteins requires the use of detergents because of the nature of the attractions holding the proteins in the membrane. Integral membrane proteins have hydrophilic exteriors that are attracted to the polar head groups of membrane phospholipids and hydrophobic interiors that interact significantly with the fatty acid tails of the membrane lipids via hydrophobic interactions (the mutual exclusion of water). Ions cannot diffuse into the membrane, and even if they could, a high salt concentration would only serve to make the hydrophobic interactions between the protein and lipid even stronger. A detergent, however, is amphipathic, and thus contains a hydrophobic and a hydrophilic portion; this allows the detergent to coax the integral protein out of the membrane. Peripheral membrane proteins, on the other hand, can be isolated using high salt concentrations. These proteins are attached to the membrane by polar interactions (like hydrogen bonding) with polar regions of integral proteins or with the polar head groups of the membrane phospholipids. When salt is added, the ions of the salt can form electrostatic/hydrogen bonds with the peripheral protein, taking the place of the attractions holding it to the membrane, and thus coaxing it away from the membrane surface.

After a man infected with the bacterium E. coli was treated with the correct antibiotic for this pathogen, the bacterium was no longer found in the man's blood, but his symptoms of fever and inflammation worsened. What caused the man's response to the treatment? Why was his condition worsened by the treatment?

The man's response to the treatment was most likely caused by the presence of the lipid A part of Lipopolysaccharides that arose from the bacterium. Lipopolysaccharides are in the outer membrane of gram-negative bacteria and can cause symptoms such as a form of septic shock if they enter the bloodstream. The medication, rather than just killing the E. coli, must have burst open the outer membrane and released these endotoxins into the bloodstream that actually worsened the man's conditions rather than improved.

*What is a nucleoid? What is the bacterial chromosome? What is a plasmid?

The nucleoid is the region of the cytoplasm of a bacterial cell that contains the genetic material and several proteins, as bacteria do not have a true membrane-bound nucleus. Most bacteria contain only one circular double-stranded chromosome. This chromosome is very tightly packaged via supercoiling with the help of nucleoid-associated proteins. While the chromosome houses genes necessary for the day-to-day survival and growth of the bacterium, many bacteria also have extrachromosomal double-stranded DNA molecules called plasmids, most of which are circular. Plasmids use the DNA-replicating machinery of the cell to replicate independently of the cell cycle. The few genes that reside on plasmids are not necessary for everyday survival and growth but often confer an advantage to the bacterium when under stress or when exposed to novel conditions in its environment. For example, a plasmid may give a bacterium antibiotic resistance or allow it to produce substances called bacteriocins that destroy closely related species.

Consumers are advised to avoid stuffing a turkey the night before cooking, even though the turkey is refrigerated. A homemaker questions this advice and points out that the bacteria of human disease growly mainly in warm temperatures, not in the refrigerator. What explanations might you offer to counter this argument?

Turkeys should not be stuffed the night before because the stuffing is moist + an uncooked turkey can lead to the growth of bacteria, such as salmonella. Stuffing the turkey the night before allows bacteria ample time to multiply until the center of the stuffing reaches refrigerator temperatures.

Viable place counts are used to estimate population size when the density of microorganisms is high, whereas membrane filtration is used when the density is low. Why is a viable plate count appropriate when the density is high, but not when the density is low?

Viable plate count only counts the cells that are able to reproduce when cultured, or the cells that were able to form a colony. It is often measured in colony forming units(CFU) rather than the number of cells. This is because it is not a known whether only one cell went into the creation of a single colony. This way of counting would cause much more inaccuracy when the density is low rather than when the density is high. Membrane filtration, on the other hand, filters out the bacteria and then grows them in a media until each cell forms a separate colony. This method would give a much more accurate reading on the cell count when the density is low.

If tryptophan is added to a culture of E. coli, further synthesis of the amino acid stops immediately. Is that a consequence of regulation of gene expression or feedback inhibition? Why?

When tryptophan is added to a culture of E. coli, further synthesis of amino acid stops immediately more than likely due to feedback inhibition. The desired concentration of amino acid may have been reached (or excess) therefore inhibiting the regulatory enzyme synthesizing the amino acids. Which in turn is controlling the overall amount of product formed. The pacemaker enzyme (which is the regulatory enzyme) in usually one of the first enzymes in the biosynthetic pathway and is regulated in feedback inhibition by being inhibited when the pathway end product is balanced for that particular case that way the organism is not making excess of whatever is being biosynthesized. I think the word "immediately" is a hint that this is feedback inhibition. Regulation by control of gene expression is a much slower process that does not manifest itself for a little while. For one, enzymes necessary to catalyze the synthesis are already present in the cell, and would stay around for a little while. With feedback inhibition, the excess tryptophan can act immediately on the regulatory enzyme and the effect is seen very quickly.

Your lab partner maintains that organisms use proteins to synthesize enzymes. You counter that organisms use enzymes to synthesize proteins. Who is right? Why?

While enzymes play a crucial role in catalyzing chemical reactions, most would not be present without proteins. There first needs to be protein synthesis, so that enzymes can assemble and then proceed to speed up other chemical reactions. However, there are ribozymes, which are made of RNA and capable of catalyzing some chemical reactions. Ribozymes are involved with cleavage or ligation of RNA and DNA, and also peptide bond formation.

If you transferred bacteria to a fresh medium and they began to grow without a lag, what would you conclude?

Without a lag phase, the bacteria increase in exponential growth by dividing rapidly. I would probably conclude that the environment was a similar medium to the previous environment and/or the microbes were not harmed in the transfer to the new medium so they do not need any "recovery" time to create new nutrients.

How do yeast cells make alcohol and cause bread to rise?

Yeast cells converts sugars of the bread and convert it into ethanol, carbon dioxide and energy. Firstly, one glucose molecule(C6H12O6) is converted into two molecules of the two carbon sugar: pyruvate (C3H4O3) through a pathway known as glycolysis. These pyruvate molecules are then oxidized to acetaldehyde(CH3CHO) and carbon dioxide by the enzyme pyruvate decarboxylase. Alcohol dehydrogenase then reduced acetaldehyde into ethanol (C2HOH).The carbon dioxide produced as a byproduct, is entrapped within the bubble and causes the dough to expand or inflate.