Exam 2 Study Guide (Midterm) PDF

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Exam 2 Study Guide (Midterm). Answers to Study guide questions given by professor....

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Chapter 5 1. Accurate definitions of disinfecting, degerming, sanitizing, etc. Disinfecting: Removal of vegetative pathogens from inanimate objects. Degerming: Mechanical removal of microbes from a limited area of live tissue (alcohol swab of skin before injection). Sanitizing: Lowering microbial counts on eating utensils to prevent transmission of pathogenic microbes. Antisepsis: Removal of pathogens from living tissue (hand washing/scrubbing using iodinated soap before surgery) Sterilization: Removal of all microbial life including endospores and viruses. Commercial Sterilization: Sufficient sterilization to killing  C. botulinum endospores in canned food. Biocide/Germicide/fungicide/virucide: Chemicals capable of killing the respective microorganisms. 2. Effect of pressure on the boiling point of water, how it affects cooking and autoclaving - Boiling point of water is higher at lower altitude (lower altitude means higher atmospheric pressure) - Pressure 14.7 PSI: boiling point 100°C - Pressure 15 PSI: boiling point 121°C - High altitude, low pressure (require more heat to cook) - Water boiled at high pressure kills bacteria more efficiently - Autoclaving- Steam heat at 15 PSI (i.e. 121°C), usually kills all microbes and spores (treatment time may be increased if the volume is large) *121  C, 15 psi, 15 min* 3. Proper processing of items before autoclaving - Use liquid cycle for autoclave liquid and dry cycle for autoclave solid objects. Close door appropriately. - Put autoclave tapes or spore indicator when autoclaving Liquid cycle (for liquid) - Never tightly close containers, allow moisture to penetrate easily - Has three phases, I.conditioning, II.exposure and III.exhaust. The autoclaving time is the time of exposure time (not the whole period) - Increase exposure time if there is a very large volume - Remove liquid when exhaust cycle is over (to prevent excess liquid loss due to evaporation)

Dry cycle (for solid objects) • Cover objects with breathable paper or cloth,do not cover with aluminum foil (check if the objects are heat stable or autoclavable). • May opt. to Keep in the autoclave even after exhaust cycle(to dry the objects). • Ensure total removal of exhaust vapor before opening the autoclave (to prevent personal injury). 4. How filtration works, limitation of filter-sterilization Filtration: A filter retains microbes while letting suspending fluid or air pass through small holes. - Does not kill microbes. Only removes them. 5. How gamma and X irradiation act in inactivating microbes (pg 127, 5.4) a. Gamma and X-ray irradiation are two forms of ionizing radiation. A gamma ray is a type of electromagnetic radiation. Gamma rays have very high frequency and very short wavelength. The gamma rays break DNA and RNA strands therefore killing the microbe. Gamma rays and X rays are both Ionizing radiation. Ionizing radiation has enough energy to remove electrons from atoms. This harms cells directly by destroying DNA and damaging cytoplasmic membranes. Gram negative bacteria is especially susceptible to this type of radiation. 6. How UV irradiation act in inactivating microbes (pg. 127, 5.4) Ultraviolet light in wavelengths of approximately 220–300 nm destroys microbes by damaging their DNA. Actively multiplying organisms are the most easily killed, whereas bacterial endospores are the most UV-resistant. 7. Limitations of UV as a controlling agent of microbes It penetrates poorly, however [UV radiation], which limits its use. Even a thin film of grease on the UV bulb or material covering microbial cells can make it less effective. Likewise, it cannot be used to destroy microbes in solid substances or turbid liquids. Because most types of glass and plastic screen out ultraviolet radiation, UV light is most effective when used at close range against exposed microorganisms. 8. Utility of freezing and flash-freezing in reducing microbe counts Although drying stops microbial growth, it does not reliably kill bacteria and fungi in or on foods. Kills all helminthes, their eggs and larvae. 9. How phenol and phenolics are related and how they work on microbes (pg 132)

● Phenolics are derivatives of phenol that have greater germicidal activity. Phenolics destroy cytoplasmic membranes of microorganisms and denature proteins. They kill most vegetative bacteria and, in high concentrations (from 5–19%), many can kill Mycobacterium species. ● Phenol was one of the first disinfectants. 10. Difference between tincture, iodine and iodophors and how iodine works on microbes (pg 131) -Iodine: mode of action unknown -Tincture: 2% solution of I and Na in alcohol, iodine-alcohol mixture, very effective in decontaminating body surfaces, used in hospitals for hand washing -Iodophors: iodine linked to organic detergent molecules 11. Why diluted alcohol is better than pure alcohol in killing microbes, how alcohols works on microbes (pg 129) - Alcohol dissolves  membranes, coagulates  proteins and dehydrates. Can be used on live tissues, effective than vegetative cells but not on spores and many fungi. - Diluted alcohol works better because denaturation of protein requires water 12. How to inactivate the prion agents - Prion agents are removed by boiling contaminated objects in 4M NaOH solution for several hours 13. How glutaraldehyde, formaldehyde, ethylene oxide and propylene oxide work, why they are not widely used (pg 129) - Glutaraldehyde - can be used to fix tissues, also a good disinfectant - Formaldehyde- formalin is used in preserving biological specimens, alkylates macromolecules, oxidizing agent - Not widely used due to toxicity and being a carcinogen - Ethylene oxide is an o rganic compound with the formula C2H4O. It is a cyclic ether and the simplest e poxide: a three-membered ring consisting of one oxygen atom and two carbon atoms. Ethylene oxide is a colorless and flammable gas with a faintly sweet odor. - Propylene oxide is an organic compound with the molecular formula CH3CHCH2O. This colourless volatile liquid with and odour resembling ether, is produced on a large scale industrially, its major application being its use for the production of polyether polyols for use in making p olyurethane plastics. It is a chiral epoxide, although it is commonly used as a r acemic mixture. 14. Difference between soaps and detergents and how they work in cleaning

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Soap itself does not destroy many organisms. It simply aids in the mechanical removal of microbes, including most pathogens, as well as dirt, organic material, and some skin cells. Soaps are made with materials found in nature. Detergents are synthetic and can be hazardous to the environment and a person's health

15. List of the common chemical sterilants and how they are used - Hydrogen peroxide- a good surface disinfectant, catalase positive bacteria may survive, used in decontaminating food container - Benzoyl peroxide- wound cleaning, used as an over the counter acne medicine - Ozone- to disinfect water and fruit juice - Peracetic acid- a very effective antimicrobial liquid, used alone (0.2%) or with hydrogen peroxide (irritant, oxidizing)

Chapter 6: Metabolism: Catabolic reactions/cellular respiration (15 questions) 1. Define metabolism, anabolism and catabolism and why they are interconnected Metabolism- the sum of the chemical reactions in an organism Anabolism - involves energy-using reactions. Larger molecules are synthesized from atoms and smaller molecules using energy from hydrolyzing ATP. Absorbs energy. Catabolism - involves energy-releasing reactions. Larger molecules are broken into smaller molecules by the reactions and the bond energy is converted to heat and bond energy of ATP. Release energy. 2. Different types of enzymes and why enzymes are important for chemical reactions of the cell (pg 147) -Oxidoreductases,Transferases, Hydrolases, Lyases, Isomerases, and Ligase - Enzymes reduces the amount of activation energy needed to start a chemical reaction. -Speeds up a chemical reaction without being used in the process. -Each enzyme acts on specific metabolic substances called substrates 3. Active sites and allosteric sites of enzymes and very basics of enzyme regulation (pg 147) - Substrates bind to the active  site, the enzyme changes shape with the substrates and creates a enzyme-substrate complex. A new product is formed and released from the enzyme. Enzyme goes back to original shape.

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Molecules bind to the allosteric sites, changing the shape of the enzyme. This can increase or decrease the affinity for the substrate.

4. Effects of temperature and pH changes on enzyme activities - Temperature: As temperature increases, so does the rate of reaction. If it gets too high, then it can denature the proteins. Starts to denature around 37 degrees C. If temperature decreases is slows the rate of reaction. - pH: Optimum pH is 7. The enzymes rate of reaction is highest at 7 and slows down the farther away from seven it goes (whether up or down). 5. What is substrate level phosphorylation and how it is used in synthesizing ATP - metabolic reaction that results in the formation of ATP or GTP by the direct transfer of a phosphoryl (PO3) group to ADP or GDP from another phosphorylated compound. 6. What you mean by substrates, reactants, and products of a chemical reaction or an enzyme-catalyzed reaction Substrate - a reactant acted upon by an enzyme - bind to active site. Reactant - the starting materials of a reaction and will always be on the left side of the equation Product - always on the right side 7. The reactants and products of glycolysis a. Glycolysis is Glucose to Pyruvate b. 2 ATP put in c. Output of 4 ATP + 2 NADH 8. The reactants and products of pyruvate dehydrogenase reaction a. Releases CO2 and 2 NADH b. Product = Acetyl CoA 9. The reactants and products of Krebs cycle a. All told, the Krebs cycle forms (per two molecules of pyruvic acid) two ATP molecules, ten NADH molecules, and two FADH2 molecules. The NADH and the FADH2 will be used in the electron transport system. Krebs cycle is a cyclic chain of enzyme-catalyzed reactions where the two carbon atoms of the acetate are oxidized and more ATP are produced The reactions take place in bacterial cytoplasm or mitochondrial matrix

During the process, two molecules of CO2, 3 molecules of NADH (=9 ATP), one molecule of FADH2 (=2 ATP) and one molecule of ATP is produced from each molecule of acetyl Co-A The NADH and FADH2 produced must be oxidized in the presence of O2 for ATP to be produced (see later) The potential net ATP production in the Kreb cycle is 12x2=24 ATP 10. When CO2 is generated and O2 is used in cellular respiration Cellular respiration is oxidation of organic molecules to produce ATP Catabolic reactions produce electrons, the electrons must be accepted by another atom 10. Definition of aerobic, anaerobic, facultative anaerobic, microaerophilic, capnophilic and fermentative microbes Aerobic respiration: when Oxygen serves as the electron acceptor(as a result H2O is produced) The overall reaction of carbohydrate catabolism is C6H12O6+6O2+36-38ADP-------->6CO2+6H2O+36-38ATP Anaerobic respiration: when organic or inorganic molecules other than O2 serves as the electron acceptor (may involve production of organic or inorganic chemicals such as lactic acid, ethanol, CO2, H2S, Fe etc.)

11. The roles of NAD, NADH, FADH, FADH2, NADP, Coenzyme A and coenzyme Q in cellular respiration Cellular Respiration consists of: Glycolysis, The Krebs cycle, and the Electron Transport Chain Coenzyme Q - small organic molecule, accepts the electron and moves it across the membrane to give it to the next electron acceptor. The cytochromes and Co-Q together form ETC Coenzyme A - Pyruvate removes Carbon to become acetic acid. Coenzyme A is added to acetic acid As a result an activated two carbon molecule (acetyl coA) and one molecule of NADH (=3 ATP) are produced from each molecule of pyruvate. Potential net ATP production at this step is 6 ATP NADP NADH and FADH2 - they are produced from Kreb’s cycle must be oxidised for ATP in the ETC. Electrons brought by NADH are transported through a series of cytochromes and Coenzyme Q. Electrons carried by FADH2 are given to Co-Q and then to the ETC 12. How ATP is synthesized by ATP synthase (pg 159) - The enzyme ATP synthase uses the energy of proton motive force to synthesize ATP. It does this by allowing protons to flow back into the bacterial cell (or matrix of the mitochondrion) in a controlled manner, simultaneously using the energy released to add a phosphate group to ADP. One molecule of ATP is formed from the entry of approximately three protons. 13. How ATP is synthesized from digesting lipids (very basic information) - Lipids are digested in the intestine by lipase. Fatty acids and glycerol are released and glycerol enters the glycolytic pathway. Fatty acids are oxidized into Acetyl CoA and Acetyl CoA enters the Kreb cycle - which produces ATP. 14. How ATP is synthesized from digesting proteins (very basic information) - Proteins are digested into amino acids by digestive enzymes (such as trypsin) and then converted into organic acids present in the Kreb cycle. Kreb cycle = ATP 15. What is fermentation and why cells ferment - Fermentation is a metabolic process where an organism converts a carb, such as a starch or sugar, into alcohol or acid, in the absence of O2. - Fermentation makes it possible to generate ATP in the absence of O2. 16. Some fermented products used by humans - Pickled vegetables, milk products (cheese, yogurt), beer, wine

Chapter 7: Macromolecular synthesis: 15 questions 1. Define genomes, the genome of a bacterium, a eukaryotic cell, a DNA virus and a RNA virus ● Genome: all DNA sequences present in a cell is called a genome ● The Genome contains genes and regulatory regions ● Prokaryotes: genes are polycistronic, not split. Simple promoters. Only one RNA polymerase. RNA polymerase itself recognizes the promoter. mRNA is used before transcription is terminated. mRNA is not modified. ● Eukaryotes: genes are monocistronic but split. Promoters are complex in structure. 3 RNA polymerases to transcribe different genes. General transcription factors recognize the promoters. RNA must be processed and transported to the cytoplasm before it can be translated. mRNA is capped and contains a polyA tail. ● The roles of DNA polymerase, DNA ligase, Primase, and Helicase, in DNA replication ● DNA polymerase- to make DNA from nucleotides, the building blocks of DNA. ● DNA ligase- an enzyme that repairs irregularities or breaks in the backbone of double-stranded DNA molecules. ● Primase- produces RNA molecules. Functions by synthesizing short RNA sequences that are complementary to a single-stranded piece of DNA. ● Helicase- continues to unwind the DNA forming the replication fork. 2. The roles of Topoisomerases, single strand DNA binding proteins and origin binding proteins in DNA replication ● Topoisomerases- enzymes that participate in the overwinding or underwinding of DNA. ● Single strand DNA binding proteins - binds to single-stranded DNA and prevents annealing of single-stranded DNA into double-stranded DNA. 3. Why DNA replication needs both ATP, CTP, GTP, UTP; and dATP, dGTP, dCTP and dTTP ● DNA primase (RNA polymerase) synthesize a small piece of RNA using ATP, CTP, GTP, UTP on one of the SS DNA strand. DNA primase then falls off. ● DNA polymerase then enters at the fork and elongates the RNA molecules using dATP, dCTP, dTTP and dGTP. 4. How the antibiotics ciprofloxacin, tetracycline, rifamycin, chloramphenicol, and neomycin act ● Ciprofloxacin- kills bacteria by stopping a bacterial enzyme called DNA-gyrase from working. Inactivates bacterial DNA topoisomerases.

● Tetracycline- By binding specifically to the 30S ribosome of the bacteria, preventing attachment of the aminoacyl tRNA to the RNA-ribosome complex. All while inhibiting other steps of the protein biosynthesis. ● Rifamycin - kills the bacteria that are causing the infection by targeting and inactivating a bacterial enzyme called RNA-polymerase. ● Chloramphenicol- it inhibits protein synthesis. It prevents protein chain elongation by inhibiting the peptidyl transferase activity of the bacterial ribosome and binds to A2451 and A2452 residues in the 23S rRNA of the 50S ribosomal subunit, preventing peptide bond formation. ● Neomycin - kills bacteria as a result of abnormal protein production in the bacterial cell it cannot produce any of its proteins correctly - and also as a result of damage to the bacterial cell. 5. Define the promoter, operator, terminator, upstream control element of a bacterial gene ● Promoter - a binding site in a DNA chain at which RNA polymerase binds to initiate transcription of messenger RNA by one or more nearby structural genes. ● Operator- a segment of DNA to which a transcription factor binds to regulate gene expression by repressing it. ● Terminator- A section of nucleic acid sequence that marks the end of a gene or operon in genomic DNA during transcription. ● Upstream control element of a bacterial gene - increases the level of gene transcription 6. Differences between the structures of a simple bacterial gene and a bacterial operon - An operon contains several genes under the control of one promoter 7. Define the template strand, top strand, coding strand and bottom strand of a gene ● Define the template strand- the strand used by DNA polymerase or RNA polymerase to attach complementary bases during DNA replication or RNA transcription. ● Top strand- 5’->3’ ● Coding strand- the DNA strand whose base sequence corresponds to the base sequence of the RNA transcript produced. ● Bottom strand of a gene - 3’->5’ 8. Explain the structural components of a bacterial mRNA Messenger RNA (mRNA) serves as the intermediary between DNA and the synthesis of protein products during translation.

It carries the genetic information copied from D  NA in the form of a series of three-b ase code “words,” each of which specifies a particular a mino acid. 9. The codons and anticodons and where are they located A codon is a three-base sequence (three nitrogen bases in a row) on mRNA. It calls for a specific amino acid to be brought to the growing polypeptide. An anticodon is a three-base sequence on tRNA. It matches the codon. 10. How to translate the ORF of a mRNA using the genetic code table

11. How to find identify the anticodon for a codon ● The codons- a sequence of three nucleotides that together form a unit of genetic code in DNA or RNA molecule. Found on molecules of tRNA. ● Anticodons- a sequence of three nucleotides forming a unit of genetic code in a transfer RNA molecule, corresponding to a complementary codon in messenger RNA. Found on molecules of tRNA. 12. Structure of ribosomes, identify the 30S, 50S, 70S, 40S, 60S and 80S ribosomes ● Bacterial cells contain 30s and 50s ribosomes, the particles remain free in the cytoplasm ○ Together 30s and 50s units make a 70s ribosome Eukaryotic cells contain 40s and 60s ribosomes and can be found bound to the exterior of the endoplasmic reticulum

○ Together 40s and 60s units make a 80s ribosome 13. Structure of tRNAs, how tRNAs are charged, what is charging ● tRNA is a short, partially DS-RNA that is clover shaped. ● The stem of the clover leaf carries an amino acid when charged by the Amino-acyl tRNA tRNA synthetase (AATase) enzymes.

14. Structure of a translation initiation complex, identify the components of the complex

Chapter 8: Microbial Genetics: 11 questions 1. Define the four types of point mutation and identify them in examples (look in ch. 8 slides) - Point mutation is a mutation that only affects a single nucleotide. - Missense mutation: a different amino acid in the protein - Read through mutation: no stop signal, a longer protein is made. - Nonsense mutation: stop signal in the middle of the gene, a very short protein is made. - Insertion/deletion mutation (frameshift): the ORF is shifted and destroyed 2. Differences between DNA replication errors and DNA damage ● Replication errors, mostly removed by m...