Cells & Biomolecules
Glycosaminoglycan (GAGs)
- A group of highly acidic polysaccharides with the structure of A-B-A-B-, where A and B represent two different sugars. The best-studied example of this is heparin, which is secreted in the lungs and tissues as a response to tissue damage. Heparin inhibits the formation of blood coagulation by activating an inhibitor (Antithrombin) of a key enzyme (thrombin), which is required for the coagulation. This response is used to ensure clots don't block the flow of blood, which is necessary to the lungs and heart, or death would inevitably ensue.
Disaccharides
- Molecules composed of only two sugar units. These serve primarily as readily available energy stores. Sucrose is a major component of plant sap, which carries chemical energy from one part of the plant to another. Lactose supplies newborns with fuel for early growth and development and is hydrolyzed by the enzyme lactase which is present in the plasma membranes of the cells that line the intestine.
Ribozymes
A RNA molecule that functions as a catalyst in cellular reactions
Atomic Force Microscope
A high resolution scanning instrument that is becoming increasingly important in nanotechnology and molecular biology. The AFM operates by scanning a delicate probe over the surface of the specimen- it can record and play the activities in real time.
Nucleus
A membrane‐bound organelle in eukaryotic cells functioning to maintain the integrity of the genetic material and, through the expression of that material, controlling and regulating cellular activities.
Protoplast
A naked plant cell whose cell wall has been digested away by the enzyme cellulase.
Guanosine Triphosphate (GTP)
A nucleotide of great importance in cellular activities. It binds to a variety of proteins (called G proteins) and acts as a switch to turn on their activities.
Polysaccharide -
A polymer of sugar unites joined by Glycosidic bonds. Glycogen is an example of a polysaccharide
a. What is a radioisotope? b. How are radioisotopes used in cell and molecular biology?
A radioisotope is a radioactive version of a molecule. It can act as a tracer, revealing its location by a radioactive signal. They are used to localize or monitor changes during an experiment.
Differential centrifugation
A technique used to isolate a particular organelle in bulk quantity, which depends on the principle that, as long as they are more dense than the surrounding medium, particles of different size and shape travel towards the bottom of a centrifuge at different rates when placed in a centrifugal field.
SDS - PAGE
A very common method for separating proteins by electrophoresis uses a discontinuous polyacrylamide gel as a support medium and sodium dodecyl sulfate (SDS) to denature the proteins. The method is called sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Polyacrylamide gel electrophoresis (PAGE), describes a technique widely used in biochemistry, forensics, genetics, molecular biology and biotechnology to separate biological macromolecules, usually proteins or nucleic acids, according to their electrophoretic mobility. Mobility is a function of the length, conformation and charge of the molecule. As with all forms of gel electrophoresis, molecules may be run in their native state, preserving the molecules' higher-order structure, or a chemical denaturant may be added to remove this structure and turn the molecule into an unstructured linear chain whose mobility depends only on its length and mass-to-charge ratio. For nucleic acids, urea is the most commonly used denaturant. For proteins, sodium dodecyl sulfate (SDS) is an anionic detergent applied to protein samples to linearize proteins and to impart a negative charge to linearized proteins. This procedure is called SDS-PAGE. In most proteins, the binding of SDS to the polypeptide chain imparts an even distribution of charge per unit mass, thereby resulting in a fractionation by approximate size during electrophoresis. Proteins that have a greater hydrophobic content, for instance many membrane proteins, and those that interact with surfactants in their native environment, are intrinsically harder to treat accurately using this method, due to the greater variability in the ratio of bound SDS.[1]
Affinity Chromatography
Affinity chromatography is a method of separating biochemical mixtures based on a highly specific interaction such as that between antigen and antibody, enzyme and substrate, or receptor and ligand. Affinity chromatography can be used to: Purify and concentrate a substance from a mixture into a buffering solution Reduce the amount of a substance in a mixture Discern what biological compounds bind to a particular substance Purify and concentrate an enzyme solution.
Posttranslational modifications (PTMs)
Alterations to the side chains of the 20 basic amino acids after their incorporation into a polypeptide chain.
Embryonic Stem Cells
An undifferentiated cell, taken from an embryo that has potential to give rise to various other cell or tissue types
Organelles
Any of several membrane-enclosed structures with specialized functions, suspended in the cytosol of eukaryotic cells.
Nucleoid
Area in prokaryotic cells in which DNA is concentrated, though not bounded by a membrane
How are proteins separated in polyacrylamide gel electrophoresis?
By their molecular weight
The chemistry of life is centered on:
Carbon
Eukaryotic
Cell characterized by an internal structure based on organelles such as the nucleus. (eukaryon: true nucleus)
Cell Line
Cells that are commonly used in tissue culture studies that have undergone genetic modifications that allow them to be grown indefinitely
Stem Cells
Cells that divide and remain undifferentiated. Three types are totipotent, pluripotent, and multi-potent.
a. What is the secondary conformation of proteins? b. What types of secondary conformations are found in proteins? c. Give an example of the property of these secondary conformations.
Conformation of portions of the polypeptide chain α-helix and β-pleated sheets An α-helix can be extended in length if subjected to pulling forces (in wool, for example), whereas β-strands are highly extended and resist pulling (in silk, for example).
disulfide bridge
Cysteine contains a reactive sulfhydryl group and is often covalently linked to another cysteine residue as a disulfide bridge: forms between two cysteines that are distance from one another in the polypeptide backbone or in two separate polypeptides. They help stabilize the intricate shapes of proteins.
Lysis:
Destruction. Hemolysis is the destruction of red blood cells with the release of hemoglobin
Ultracentrifugation
Differential centrifugation is a common procedure in microbiology and cytology used to separate certain organelles from whole cells for further analysis of specific parts of cells. In the process, a tissue sample is first lysed to break the cell membranes and mix up the cell contents. The lysate is then subjected to repeated centrifugations, each time removing the pellet and increasing the centrifugal force. Finally, purification may be done through equilibrium sedimentation, and the desired layer is extracted for further analysis. Separation is based on size and density, with larger and denser particles pelleting at lower centrifugal forces. As an example, unbroken whole cells will pellet at low speeds and short intervals such as 1,000g for 5 minutes. Smaller cell fragments and organelles remain in the supernatant and require more force and greater times to pellet. In general, one can enrich for the following cell components, in the separating order in actual application: Whole cells and nuclei; Mitochondria, chloroplasts, lysosomes, and peroxisomes; Microsomes (vesicles of disrupted endoplasmic reticulum); and Ribosomes and cytosol.
Polypeptide Chains
During protein synthesis the joining of AA's forming long unbranched polymers is called a polypeptide chain.
Electrophoresis
Electrophoresis" refers to the electromotive force (EMF) that is used to move the molecules through the gel matrix. By placing the molecules in wells in the gel and applying an electric field, the molecules will move through the matrix at different rates, determined largely by their mass when the charge to mass ratio (Z) of all species is uniform. However, when charges are not all uniform then, the electrical field generated by the electrophoresis procedure will affect the species that have different charges and therefore will attract the species according to their charges being the opposite. Species that are positively charged will migrate towards the cathode which is negatively charged (because this is an electrolytic rather than galvanic cell). If the species are negatively charged they will migrate towards the positively charged anode.[4]
Two of the most common linkages between different functional groups are:
Ester Bonds - The Chemical bond that forms between carboxylic acids and alcohols (or acidic and alcoholic functional groups) while producing a molecule of water. Amide Bonds - The chemical bond that forms between carboxylic acids and amines (or acidic and amino functional groups) while producing a molecule of water.
Name three types of lipid molecules and state the role of each.
Fats are very rich in chemical energy. Their function is energy storage. Steroids are components of animal cell membranes and precursors of steroid hormones. Cholesterol is a steroid involved in membrane fluidity. Phospholipids function primarily in cell membranes.
Unsaturated & Saturated & Oils
Fatty acids typically vary in length from 14 to 20 carbons. Those that lack double bonds are described as saturated. Those possessing double bonds are unsaturated. The more double bonds a fatty acid contains the less effectively these chains can be packed together, which in turn lowers its melting temperature. Fats that are liquid at room temperature are labeled oils. Solid formed foods made from oil (margarine) are created through hydrogenation. This process converts the cis bonds into trans bonds, which are straight instead of kinked. Fats are very rich in chemical energy; one gram of fat contains over twice the energy content of a gram of carbohydrate. Carbs function as a short-term rapidly available energy source, whereas fat reserves store energy on a long-term basis. FUN FACT the average person contains about 0.5 kg's of carb storage primarily in the form of glycogen (2000kcal of energy) the average person also contains roughly 16 kg's of fat (144,000 kcal of energy). Due to the lack of polar groups fat in very insoluble in water therefore is stored in cells in the form of dry lipid droplets. In many animals fat is stored in special cells called adipocytes, which exhibit the ability to change volume according to varying quantities of fat.
a. What is fluorescence microscopy? b. What is immunofluorescence? What is it used for?
Fluorescence microscopy is a technique that uses certain fluorescent compounds to stain specimens. In immunofluorescence, a fluorochrome is linked to an antibody to produce a fluorescent antibody. It is used to determine the location of a specific protein in the cell.
Name three polysaccharides composed of polymers of glucose.
Glycogen, starch, and cellulose Glycogen is found in animal cells, whereas starch is found in plant cells. Cellulose is the main component of plant cell walls.
Hematopoietic stem cells
Hematopoietic stem cells (HSCs) are multipotent, self-renewing progenitor cells that develop from mesodermal hemangioblast cells. All differentiated blood cells from the lymphoid and myeloid lineages arise from HSCs. HSCs can be found in adult bone marrow, peripheral blood, and umbilical cord blood.
Glycogen -
Highly branched glucose polymer that serves as a readily available chemical energy in most animal cells. Glycogen is an insoluble polymer of glucose, which is stored in the tissues of the liver. Various food materials are carried to the liver where they are chemically converted to glucose and stored as glycogen. Human skeletal muscles typically store enough glycogen to fuel about 30 minutes of moderate activity.
a. Distinguish between homogenization and cell fractionation. b. How do you separate purified cellular components after fractionation?
Homogenization is the breaking of cells by mechanical disruption, whereas cell fractionation is the purification of specific parts of a homogenate, usually containing organelles. Organelles are purified by differential centrifugation. For details, see Figure 18.23.
There are two types of basic viral infections:
In most cases the virus stops normal cell activity and redirects the cells materials to manufacture viral nucleic acids and proteins, which assemble into more virions which are eventually released when the infected cell ruptures and these virions can then infect neighbouring cells. In other cases the infected cell does not lead to the death of the infected cell but rather the virus inserts its DNA into the DNA of the hosts cells chromosomes. The integrated DNA is called a Provirus. Proviruses have different effects depending on the viruses. One outcome is the cells containing the provirus behave normally until exposed to a stimulus like ultraviolet radiation-which activates the dormant cells leading to the lysis of the cell and the release of viral progeny. The other outcome is the provirus infected cells produce new viral progeny that bud at the cells surface without lysing the infected cell- this allows the host cell to remain alive for a period of time acting like a factory for the production of new virions.
Gel Electrophoresis
In simple terms, electrophoresis is a process which enables the sorting of molecules based on size. Using an electric field, molecules (such as DNA) can be made to move through a gel made of agar or polyacrylamide. The electric field consists of a negative charge at one end which pushes the molecules through the gel, and a positive charge at the other end that pulls the molecules through the gel. The molecules being sorted are dispensed into a well in the gel material. The gel is placed in an electrophoresis chamber, which is then connected to a power source. When the electric current is applied, the larger molecules move more slowly through the gel while the smaller molecules move faster. The different sized molecules form distinct bands on the gel.[citation needed] The term "gel" in this instance refers to the matrix used to contain, then separate the target molecules. In most cases, the gel is a crosslinked polymer whose composition and porosity is chosen based on the specific weight and composition of the target to be analyzed. When separating proteins or small nucleic acids (DNA, RNA, or oligonucleotides) the gel is usually composed of different concentrations of acrylamide and a cross-linker, producing different sized mesh networks of polyacrylamide. When separating larger nucleic acids (greater than a few hundred bases), the preferred matrix is purified agarose. In both cases, the gel forms a solid, yet porous matrix.
Whole mount
Is an intact object, either alive or dead and can consist of an entire microscopic organism or a small part of a large organism. Most tissues are too opaque unless examined as a slice.
Lipids
Lipids are a diverse group of nonpolar molecules whose common property is the ability to dissolve inorganic solvents, and their inability to dissolve in water.
Fatty Acids
Long Unbranched hydrocarbon chains with a single carboxylic acid group at one end. Fatty acids contain the inherent quality of being amphipathic due to the fact that its hydrocarbon chain is hydrophobic and its carboxyl group (-COOH, which bears a negative charge) is hydrophilic.
The building blocks of macromolecules.
Macromolecules within a cell have a short lifetime compared to the cell itself. With the exception of the cells DNA the macromolecules are continuously being broken down and replaced by new macromolecules. Due to this process, most cells have a supply of low-molecular-weight precursors ready to incorporate into macromolecules. This includes sugar polysaccharides. Amino Acids proteins. Nucleotides Nucleic Acids. Fatty Acids Lipids.
a. Distinguish between magnification and resolution. b. Why is it necessary to stain specimens to be viewed in a bright-field microscope?
Magnification produces an enlarged image of an object, whereas resolution is the ability to see two neighbouring points in the visual field as distinct entities. Specimens that have been thinly sectioned are translucent. To become visible, they need to be stained with a dye.
Mass Spectrometer
Mass spectrometry (MS) is an analytical technique that ionizes chemical species and sorts the ions based on their mass to charge ratio. In simpler terms, a mass spectrum measures the masses within a sample. Mass spectrometry is used in many different fields and is applied to pure samples as well as complex mixtures. A mass spectrum is a plot of the ion signal as a function of the mass-to-charge ratio. These spectra are used to determine the elemental or isotopic signature of a sample, the masses of particles and of molecules, and to elucidate the chemical structures of molecules, such as peptides and other chemical compounds. In a typical MS procedure, a sample, which may be solid, liquid, or gas, is ionized, for example by bombarding it with electrons. This may cause some of the sample's molecules to break into charged fragments. These ions are then separated according to their mass-to-charge ratio, typically by accelerating them and subjecting them to an electric or magnetic field: ions of the same mass-to-charge ratio will undergo the same amount of deflection.[1] The ions are detected by a mechanism capable of detecting charged particles, such as an electron multiplier. Results are displayed as spectra of the relative abundance of detected ions as a function of the mass-to-charge ratio. The atoms or molecules in the sample can be identified by correlating known masses to the identified masses or through a characteristic fragmentation pattern. Applications: 10.1 Isotope ratio MS: isotope dating and tracing 10.2 Trace gas analysis 10.3 Atom probe 10.4 Pharmacokinetics 10.5 Protein characterization 10.6 Glycan analysis 10.7 Space exploration 10.8 Respired gas monitor 10.9 Preparative mass spectrometry
Starch -
Mixture of two glucose polymers, amylose and amylopectin, that serves as readily available chemical energy is most plant cells. Starch is stored as densely packed granules or starch grains, which are enclosed, in membrane-bound organelles (plastids) within the plant cell. Animals don't synthesize starch, but they posses and enzyme (amylase) that hydrolyzes it.
Fats
Molecules consisting of a glycerol backbone linked by ester bonds to three fatty acids, also termed triacylglycerols.
Cells are typically microscopic because:
Most eukaryotic cells contain a single nucleus, which contains only two copies of most genes. Due to the fact that genes serve as template for the production of information carrying messenger RNA, a cell can only produce a limited number of these RNA messengers in a given period of time. The greater the cells cytoplasm volume, the longer it will take to synthesize the number of messages required by that cell. As a cell increases in size, the surface area/volume ratio decreases. The cells ability to exchange substances with its environment is proportional to its surface area. If it were to grow beyond a certain size, its surface would not be enough to take up the substances needed to support metabolic activities. A cell depends larger on random movement of molecules (diffusion). The time required for diffusion is proportional to the square of the distance to be transferred. Therefore if the cell becomes larger the time require for diffusion increases to a point where its metabolic activity is prohibitively long.
Nitrogen Fixation
N2 is converted to ammonia NH3. Bacteria do this (esp. Rhizobium living symbiotically with legumes). then it can be used by the cells in the synthesis of nitrogen containing organic compounds, including amino acids and nucleotides.
nucleotides DNA & RNA.
Nucleic acids are macromolecules constructed of long strands of monomers called nucleotides (composed of a five carbon sugar(ribose), a nitrogenous base, and a phosphate group). Two types of nucleic acids found in living organisms are DNA & RNA. DNA serves as the genetic material of all cellular organisms, though RNA carries out the role for many viruses. DNA governs cell activities through RNA messages.
Adenosine Triphosphate (ATP)
Nucleotide consisting of adenosine bonded to three phosphate groups; it is the principal immediate-energy source for prokaryotic and eukaryotic cells,
a. Describe the structure of nucleotides and the manner in which these monomers are strung together to form a polynucleotide strand. b. What is the evolutionary significance of ribozymes?
Nucleotides are the monomers of nucleic acids. A nucleotide consists of three parts: a sugar (ribose or deoxyribose), a phosphate group, and a nitrogenous base (a pyrimidine or a purine). Nucleotides are joined to form strands by covalent bonds that link the 3' hydroxyl group of one sugar with the 5' phosphate group of the adjoining sugar. (Refer to Figure 2.55.) Ribozymes have led to speculation that early in evolution neither DNA nor proteins existed on Earth. Instead, RNA served both as genetic material and as enzymes.
Model Organelles
Organisms that have been widely used for research so that a great deal is known about their biology. think- the fruit fly
Functional Groups
Particular groupings of atoms that tend to act as a unit, often affecting the chemical and physical behaviour of the larger organic molecules to which they belong. Functional groups often give the organic molecules their physical properties, chemical reactivity and solubility.
Phase Contrast
Phase-contrast microscopy is particularly important in biology. It reveals many cellular structures that are not visible with a simpler bright-field microscope, as exemplified in the figure. These structures were made visible to earlier microscopists by staining, but this required additional preparation and killed the cells. The phase-contrast microscope made it possible for biologists to study living cells and how they proliferate through cell division.[1] After its invention in the early 1930s,[2] phase-contrast microscopy proved to be such an advancement in microscopy, that its inventor Frits Zernike was awarded the Nobel prize (physics) in 1953
Triacylglycerol
Polymers consisting of a glycerol backbone linked by ester bonds to three fatty acids, commonly called fats.
DIfferentiation
Process in which cells become specialized in structure and function
prokaryotes have what kind of chromosome
Prokaryotes (nearly all) contain a single, circular chromosome.
specificity
Proteins exhibit a high degree of specificity: the property of selective interaction between components of a cell that is basic to life. This is due to the virtually limitless molecular structures that proteins as a group can assume. Each protein has a unique and defined structure, which enables it to carry out a particular, selective function. Proteins are polymers that are comprised of amino acids, each with its own unique sequence of amino acids. Much of the capabilities of a protein can be understood by looking at the chemical properties of its amino acids. TWENTY different amino acids are used in the construction of proteins.
Side Chains/ R groups
Side Chains/ R groups The defining functional group of an amino acid, which can range from a single hydrogen to complex polar or non-polar units in the 20 amino acids most commonly found in cells. Therefore amino acids are classified based on their side chains and fall roughly into these four categories: 1. Polar, charged side chains contain relatively strong organic acids/bases 2. Polar, Uncharged side chains are partially negative/positive & therefore form hydrogen bonds with other molecules/water. Quite Reactive 3. Nonpolar these side chains are hydrophobic & don't interact with water 4. The other three amino acids (glycine, proline, cysteine)
Single Particle Reconstruction
Single particle analysis segments and averages many particles from a sample, allowing for computer algorithms to process the individual images into a combined "representative" image. This allows for improvements in signal to noise, and can be combined with deconvolution to provide limited improvements to spatial resolution in the image.
Gel Filtration
Size-exclusion chromatography (SEC), also known as molecular sieve chromatography,[1] is a chromatographic method in which molecules in solution are separated by their size, and in some cases molecular weight.[2] It is usually applied to large molecules or macromolecular complexes such as proteins and industrial polymers. Typically, when an aqueous solution is used to transport the sample through the column, the technique is known as gel-filtration chromatography, versus the name gel permeation chromatography, which is used when an organic solvent is used as a mobile phase. The main application of gel-filtration chromatography is the fractionation of proteins and other water-soluble polymers, while gel permeation chromatography is used to analyze the molecular weight distribution of organic-soluble polymers. Either technique should not be confused with gel electrophoresis, where an electric field is used to "pull" or "push" molecules through the gel depending on their electrical charges.
Oligosaccharides
Small chains composed of sugars covalently attached to lipids and proteins; they distinguish one type of cell from another (because they come in so many different combinations of sugar units) and help mediate interactions of a cell with its surroundings. Oligos are particularly important on the glycolipids and glycoproteins of the plasma membranes where they project from the cells surface.
Prokaryotic When was the first evidence of these?
Structurally simple cells that do not have membrane bound organelles; (pro-karyon: before the nucleus) ie. archeabacteria 2.7 billion years old
In terms of the four types of macromolecules, which monomers are the building blocks of polymers?
Sugars are precursors of polysaccharides. Amino acids are precursors of proteins. Nucleotides are precursors of nucleic acids. Fatty acids are incorporated into lipids.
Freeze-Facture Replication
Technique in which a tissue sample is first frozen and then struck with a blade that fractures the tissue along the lines of least resistance, often resulting in a fracture line between the two leaflets of the lipid bilayer; metals are then deposited on the exposed surfaces to create a shadowed replica that is analyzed by electron microscopy.
Freeze etching
Techniques in which tissue is freeze fractures, then exposed briefly to a vacuum so a thin layer of ice can evaporate from above and below the fractured surfaces, exposing features for identification by electron microscopy.
Peptide Bonds
The AA's are joined together by peptide bonds which are formed through the linkage of the carboxyl group to one amino acid group with the elimination of a water molecule. Once incorporated into a polypeptide chain the AA's are termed residues. On the side of the chain with an unbonded amino group its called the N-terminus. On the side of the chain with the unbonded carboxyl group its called C- terminus.
Prokaryotes are divided into two major groups
The Archeae (or archaeabacteria) and Bacteria (or eubacteria). The Archeae group is more closely related to eukaryotes then to the eubacteria.
Isoelectric Point
The PH at which the negative charges of the component amino acids of a protein equal the positive charges of the component amino acid, so the protein is neutral.
Amphipathic
The biologically important property of a molecule having both hydrophobic and hydrophilic regions
Glycosidic Bonds
The chemical bond that forms between sugar molecules
Fluorescense
The emmision of a glowing light by a certain substances when struck by a particula wavelength (e.g. calcium tungstate screens have phosphors that emit lue light, or fluoresce, when exposed to x-ray)
Multiprotein complex
The interaction of more than one complete protein to for a larger, functional complex.
Objective Lens
The lens of a light microscope that focuses light rays from the specimen to form a real, enlarged image of the object within the column of the microscope
Limit of Resolution
The resolution attainable by a microscope is limited by the wavelength of the illumination according to the equation outlined.
What is the evolutionary importance of these common properties between prokaryotes and eukaryotes?
The similarities reflect the fact that eukaryotic cells almost certainly evolved from prokaryotic ancestors. Because of their common ancestry, the two types of cells share an identical genetic code that uses DNA and RNA, a common set of metabolic pathways, and many common structural features.
The structure of all amino acids:
The structure of amino acids all have a carboxyl group and an amino group which are separated from each other by a single carbon atom. Amino Acids also have asymmetric carbon atoms (with the exception of glycine) the carbon of amino acid bonds to four different groups so each can exist in a D or L shape FIGURE 2.25 Page 50. Amino acids used in the synthesis of proteins on a ribosome are always L-amino acids.
Why are viruses not considered to be living organisms? What are the two types of viral infections? Explain.
They are not alive. They are obligatory intracellular parasites that cannot reproduce without the presence of a host cell, animal, plant, or bacterium. Lytic infections: The virus redirects the cell to use its available materials to manufacture viral nucleic acids and proteins, which assemble into new virions. Ultimately the virus lyses or ruptures the cell and releases a new generation of viral particles capable of infecting neighbouring cells. Lysogenic infections: The infecting virus does not rupture the cell but instead inserts or integrates its DNA into the DNA of the host cell's chromosomes. This is called a provirus. It may have different effects depending on the type of virus.
Laser Scanning Confocal
Three dimensional recreations of objects
What is X-ray diffraction used for?
To determine protein and DNA structure at atomic resolution.
Distinguish between transmission and scanning electron microscopy.
Transmission electron microscopy forms images using electrons that are transmitted through a specimen, allowing the visualization of inner structure. Scanning electron microscopy forms images from electrons that bounce off the surface of the specimen, allowing the visualization of the surface of objects.
Yeast-2-hybrid
Two-hybrid screening (also known as yeast two-hybrid system or Y2H) is a molecular biology technique used to discover protein-protein interactions (PPIs)[1] and protein-DNA interactions[2][3] by testing for physical interactions (such as binding) between two proteins or a single protein and a DNA molecule, respectively. The premise behind the test is the activation of downstream reporter gene(s) by the binding of a transcription factor onto an upstream activating sequence (UAS). For two-hybrid screening, the transcription factor is split into two separate fragments, called the binding domain (BD) and activating domain (AD). The BD is the domain responsible for binding to the UAS and the AD is the domain responsible for the activation of transcription.[1][2] The Y2H is thus a protein-fragment complementation assay.
Cellulose -
Unbranched glucose polymer with (14) linkages that assembles into cables and serves as a principal structural element of plant walls. Almost all multicellular animals lack the enzyme needed to degrade cellulose, which happens to be the most abundant organic material on earth and rich in chemical energy. Animals that do digest it harbour bacteria and protozoa that synthesize the necessary enzyme cellulose. Chitin is another polysaccharide made of a different type of sugar N-acetylglucosamine. Chitin is a structural material used among invertebrates, namely the outer coverings of insects, crustaceans and spiders. Insects owe much of their success to this tough resilient plastic like material
Protists
Unicellular protists are the most complex prokaryotic cells, due to the fact that all the machinery necessary for the cells activities must be present within the confines of a single cell. The evolution of this type of cell led to multicellular organisms in which specialized cells conduct different types of activities. Differentiation is the process by which specialized cells are formed.
X-ray Crystallography
X-ray crystallography is a tool used for identifying the atomic and molecular structure of a crystal, in which the crystalline atoms cause a beam of incident X-rays to diffract into many specific directions. By measuring the angles and intensities of these diffracted beams, a crystallographer can produce a three-dimensional picture of the density of electrons within the crystal. From this electron density, the mean positions of the atoms in the crystal can be determined, as well as their chemical bonds, their disorder and various other information. The method also revealed the structure and function of many biological molecules, including vitamins, drugs, proteins and nucleic acids such as DNA. X-ray crystallography is still the chief method for characterizing the atomic structure of new materials and in discerning materials that appear similar by other experiments. X-ray crystal structures can also account for unusual electronic or elastic properties of a material, shed light on chemical interactions and processes, or serve as the basis for designing pharmaceuticals against diseases.
Protein Domains
a region within a protein that folds and functions in a semi-independent manner
a. What is the source of energy that supports life on Earth? b. How is this energy passed from one organism to the next?
a. What is the source of energy that supports life on Earth? Most of the energy required by life on Earth's surface arrives in the form of electromagnetic radiation from the sun. (Remember that some prokaryotes are chemoautotrophs.) Light-absorbing pigments present in the membranes of photosynthetic cells trap the energy that drives all cellular metabolism. b. How is this energy passed from one organism to the next? Light energy is converted by photosynthesis into chemical energy that is stored in energy-rich carbohydrates. These carbohydrates are in turn the source of energy for animal cells.
Steroids
are built around a four-ringed hydrocarbon skeleton. One of the most important steroids is cholesterol, which is responsible for the synthesis of hormones. This fat is primarily absent in plant cells.
Biofilms
are complex multispecies communities of prokaryotes.
Biochemicals
are compounds synthesized by living organisms.
Viruses
are defined as small, obligatory intracellular pathogens that are not considered to be alive because they cannot divide directly, which is required by the cell theory of life. They cannot reproduce without a host i.e. plant, animal or bacterial cell. Outside of the host, the virus exists only as a particle or Virion.
Conformational changes
are described as predictable (non-random) movements within a protein that are triggered by the binding of specific molecules. Virtually every activity in which a protein takes part is accompanied by a conformational change.
Macromolecules
are huge, highly organized carbon based atoms that form the structure and carry out the activities of a cell. These molecular giants are large in size and intricate in shape, due to this they are able to perform tasks with great precision and efficiency. Macromolecules more then any other characteristic, endow organisms with the property of life. Macromolecules can be divided into four major categories; proteins, nucleic acid, polysaccharides, and certain lipids. Proteins, nucleic acid and polysaccharides are considered polymers (composed of a large number of low-molecular-weight building blocks-monomers). These macromolecules are constructed from monomers by a process of polymerization, which resembles coupling railroad cars on a train.
Virions
are macromolecular packages that contain either DNA (double stranded) or RNA (single stranded) genetic material that is enclosed by a protein capsule called capsid. Virions are inanimate particles that cannot reproduce, metabolize or carry on any life activity. Viral capsids are made up of a specific number of subunits. If the coat is made up of many copies of the same protein then only one or a few genes are required to code for its protein container. Many viruses have a capsid whose subunits are organized into polyhedron (planar faced structure). A very popular one is the icosahedron (20 sided page 24 of text book).
Metabolic Intermediates (metabolites)
are the compounds formed along the way in the process of converting compounds A into B then into C until finally into the functional end (proteins or lipids ect.) The in between products are called metabolic intermediates.
Cyanobacteria
are the most complicated prokaryotes, which contain elaborate arrays of cytoplasmic membranes, which serve as sites of photosynthesis. These membranes are very similar to chloroplasts in plant cells. Photosynthesis in cyanobacteria is accomplished by splitting water molecules, which releases molecular oxygen. Cyanobacteria are often capable of not only photosynthesis but also nitrogen fixation (which is the conversion of nitrogen gas into reduced forms of nitrogen such as ammonia- which can then be used by the cells in the synthesis of nitrogen containing organic compounds, including amino acids and nucleotides)
Ribosomes
are the workbenches upon which proteins of the cell are manufactured. These exist in both types of cells although they have different dimensions.
Video Image Processing
beneficial when specimens are light sensitive
Van der walls
betweentwo atoms exhibiting a transient charge due to a momentary asymmetry in the distribution of electrons around the atoms.
Hydrogen
covalently bonded hydrogen groups
Purines
double ringed: guanine, Adenine
Ionic
full charged positive and negative groups
covalent bond
hold atoms together to form molecules, sharing outer electrons in a shell.
Carbohydrates (called Glycans)
include simple sugars and all larger molecules composed of sugar building blocks. Carbs function is the stores of chemical energy, and to be the durable material used for biological construction. The important sugars have the formula (CH2O)n -where n has a range from 3-7. 3 Trioses -- 4 Tetroses -- 5 Pentoses -- 6 Hexoses -- 7 Heptoses
Autoradiography
is a broad-based technique used to determine where a particular isotope is located, in a cell, in polyacrylamide gel, or on a nitrocellulose filter. Autoradiography takes advantage of the ability of a particle emitted from a radioactive atom to activate a photographic emulsion (like an x-ray).
Fixative
is a chemical solution that rapidly penetrates the cell membrane and immobilizes all of its macromolecular material so that the structure of the cell is as close to living state as possible. (formaldehyde, alcohol, acetic acid)
Chromatin
is a complex of DNA and proteins that forms chromosomes within the nucleus of eukaryotic cells. Nuclear DNA does not appear in free linear strands; it is highly condensed and wrapped around nuclear proteins in order to fit inside the nucleus.
Angstrom
is equal to one tenth of a nanometer(which is 10 to the minus nine) one angstrom is the equivalent of the diameter of a hydrogen atom. DNA is 2 nm in width
Stereoisomerism
is the arrangement of atoms in molecules whose connectivity remains the same but their arrangement in space is different in each isomer. The enzymes present in living cells can distinguish between D and L formed of a sugar. Typically only of the stereoisomers are used by the cells (such as D-glucose or L-fucose)
Metagenome
is the catalogue and sequential evaluation of virtually all the genes present in the microbes of a given habitat developing a collective genome. This approach can tell you about the types of proteins these organisms manufacture and this about many of their metabolic activities.
Human Microbiome
is the collection of trillions of unseen passengers that live on or within our own bodies. The bacteria within the intestinal tract are responsible for the synthesis of vitamins, the breakdown of complex plant sugars & the prevention of growth of pathogenic organisms.
Negative Staining
is the process by which heavy metal deposits are collected everywhere on a specimen grid except where the particles are present- the specimen then stands out by relative brightness.
Chromatography
is the term for a variety of techniques in which a mixture of dissolved components is fractionated as it moved through some type of porous matrix
Denaturation
is the unfolding or disorganization of a protein. This is brought on by a number of agents: detergents, organic solvents, radiation, heat, and compounds such as urea and guanidine chloride- all of which interfere with the various interactions tat stabilize a protein's tertiary structure.
Phase Contrast Microscope
makes a highly transparent object more visible. This type of microscope converts difference in refractive index into differences in intensity (relative brightness/darkness) made visible to the eye. This allows living cells to be examine at relatively high resolution.
Ion Exchange Chromatography
method of chromatography in which the beads in the column are coated with charged substances so that they attract or bind compounds that have opposite charge Ion chromatography (or ion-exchange chromatography) is a chromatography process that separates ions and polar molecules based on their affinity to the ion exchanger. It works on almost any kind of charged molecule—including large proteins, small nucleotides, and amino acids. It is often used in protein purification, water analysis, and quality control.[citation needed] The water-soluble and charged molecules such as proteins, amino acids, and peptides bind to moieties which is oppositely charged by forming covalent bonds to the insoluble stationary phase.[1] The equilibrated stationary phase consists of an ionizable functional group where the targeted molecules of a mixture to be separated and quantified can bind while passing through the column. This method applies the idea of the interaction between molecules and the stationary phase which are charged oppositely to each other.
Chromatography
mixture of dissolved components is fractionated through a porous matrix
Polar molecules of importance
o n s p c h
Confocal Microscope
produces an image of a thin plane situated within a much thicker specimen.
Pyrimidines
single ringed cytosine, thymine (1 extra methyl group), uracil (only in RNA)
Viroids
small obligatory intracellular pathogens that unlike viruses consist only of an uncoated circle of genetic material RNA - they are thought to cause disease by interfering with the cells normal path of gene expression.
Cytosol
soluble phase of cytoplasm
Bright field Microscope
the cone of light that illuminates the specimen is shown as a bright background against which the image of the specimen must be contrasted. This is ideally suited for specimens of high contrast, like stained tissues.
Native Self- Assembly Molecular Chaperones
the correct, or proper state. the property of proteins (or other structures) to assume the correct (native) conformation based on the chemical behavior dictated by the amino acid sequence. various families of proteins whose role is to assist the folding and assembly of proteins by preventing undesirable interactions
Contrast
the difference in appearance between adjacent parts of an object or between an object and its background.
Endosymbiont theory
the theory that mitochondria and plastids, including chloroplasts, originated as prokaryotic cells engulfed by an ancestral eukaryotic cell. The engulfed cell and its host cell then evolved into a single organism.
Phospholipids
these molecules resemble a fat but only have two fatty acid chains rather then three, making it a diacylglycerol. The third hydroxyl of the glycerol backbone is covalently bonded to a phosphate group, which is in turn covalently bonded to a small polar group such as choline. This causes phospholipids to have two distinct ends, a hydrophilic (phosphate group) end and a hydrophobic end (fatty acid tail). FIGURE 2.22 PAGE 49.
non colavent bonds
weak attractive forces between negative and positive charged regions. Ionic, hydrogen & wanderwalls
Features of Eukaryotic Cells Only:
• Division of cells into nucleus and cytoplasm, separated by a nuclear envelope containing complex pore structures • Complex chromosomes composed of DNA and associated proteins that are capable of compacting into mitotic strictures • Complex membranous cytoplasmic organelles (endoplasmic reticulum, Golgi complex, lysosomes, endosomes, peroxisomes, glyoxisomes) • Specialized cytoplasmic organelles for aerobic respiration (mitochondria) and photosynthesis (chloroplast) • Complex cytoskeletal system (including microfilaments, intermediate filaments and microtubules) and associated motor proteins • Complex flagella and cilia • Ability to ingest particulate material by enclosure within plasma membrane vesicles (phagocytosis) • Cellulose - containing cell walls (in plants) • Cell division using microtubules-containing mitotic spindle that separates chromosomes • Presence of two copies of genes per cell (Diploidy) one from each parent • Presence of three different RNA synthesizing enzymes (RNA polymerases) • Sexual reproduction requiring meiosis and fertilization
Most proteins can be categorized on the basis of their overall conformation being either:
• Fibrous proteins- one with a tertiary structure that is greatly elongated, resembling a fiber. • Globular proteins- one with a tertiary structure that is compact, resembling a globe. Most proteins that act as structural materials outside living cells are fibrous proteins (collagens, elastin's, keratin) whereas most proteins within a cell are globular. -
Common Features Between Eukaryotic & Prokaryotic Cells:
• Plasma Membrane of Similar Construction • Genetic Information Encoded in DNA using identical genetic code • Similar Mechanisms for transcription/translation of genetic info including similar ribosomes • Shared metabolic pathways (glycolysis & TCA cycle) • Similar apparatus for conservation of chemical energy as ATP (Located in the plasma membrane of prokaryotes and the mitochondrial membrane of eukaryotes) • Similar mechanisms for photosynthesis (between cyanobacteria and green plants) • Similar mechanisms for synthesizing and inserting membrane proteins • Proteasomes (protein digesting structures) of similar construction (between archaebacteria and eukaryotes)
Beta- Pleated sheet
• one possible secondary structure of a polypeptide in which several b-strands lie parallel to each other, creating the conformation of a sheet The portions of the polypeptide chain not organized into these two structures are flexible- and often the greatest site of biological activity.
Alpha helix-
• one possible secondary structure of polypeptides, in which the backbone of the chain forms a spiral conformation
