Lecture 1 In-Class Exam Explanations Pg.

186 Protein Folding Process The key point here is to realize that in the passage it explicitly states that the three dimensional shape of a protein is ultimately determined by its amino acid sequence (primary structure). The two types of proteins described assist in this process but do not directly influence it. Chaperone proteins stabilize properly folded sections of polypeptides and some enzymes catalyze reactions in the folding process allowing them to occur faster by lowering the activation energy. 1). The key point here is that enzymes catalyze chemical reactions in both directions, meaning they lower the activation required for the forward and reverse direction. This is why C. Protein disulfide isomerase increases both the rate at which disulfide bonds are formed and broken. 2). Main point: The folding pattern of a protein is determined by the protein's primary structure, the number and sequence of amino acids in the protein. 3). Which of the following is the best explanation for why attempts at predicting protein configuration based upon amino acid sequence have been unsuccessful? Since we know from the passage that the chaperone and folding process proteins only assist in the folding process and do not actually help to determine the three dimensional shape of a protein, we can cancel out answer choice B. Answer choice A is nonsensical and is not the best answer! D is also nonsensicalby definition if two proteins are the same, their amino acid sequence should be exactly the same unless a mutation has taken place. Thus, our answer is C, the three dimensional shape of a protein is based upon hydrogen and disulfide bonding between amino acids, and the number of possible combinations of bonding amino acids makes prediction difficult. You learned this from biochem. 4). The key point here is that chaperone proteins assist the folding process which determines the three dimensional shape of a protein or in other words, Tertiary Structure. Our answer is C. 5). Why would the cell increase chaperone synthesis when exposed to heat shock? A. Heat destabilizes intermolecular bonds making protein configuration more difficult to achieve. Chaperones counteract this by stabilizing the partially folded intermediates. This is the only answer choice that makes sense. The cell wants to prevent its proteins from being denatured due to the high heat. 6). Which of the following bonds in a protein is likely to be least stable in the presence of heat? The main point here is to rank the strengths of bonds found in the proteins. The strongest is the fully covalent carbonly bond, then peptide bonds, then disulfide bonds, and lastly H-bonds. Least stable in the presence of heat means which of the following is the weakest and will break first under heat? Our answer is H-bonds. 7). The key point here is which amino acid residue forms disulfide bonds? Cysteine residues form disulfide bonds.

Gel Electrophoresis This passage goes into the details of gel electrophoresis. First it talks about polyacrylamide gel electrophoresis which you performed in your lab at UCSD. The molecular basis of this technique is that one makes a gel out of polyacrylamide which then creates different sized pores. This gel is then placed in a electric field where the governing rule is run to red. Electrophoresis is weird because the positive terminal is actually the anode of the battery and the negative terminal is the cathode. So negatively charged molecule such as DNA run to the anode! Polyacrylamide is able to separate molecules based upon their size and charge ratio. Nucleic acids such as DNA already have a consistent charge/mass ratio and so it can be separated by size. In the case of proteins SDS or sodium dodecyl sulfate is added so that the proteins can be separated solely by size. The way SDS works is that it coats the protein with a uniform negative charge by denaturing the protein, disrupting its noncovalent interactions. It thereby produces proteins with a consistent charge/mass ratio. Once charge is standarzied within all of the compounds being tested, size is used to separate them. Larger molecules move more slowly through the gel because they cannot get through the pores as easily and thus usually remain higher up on the gel. Smaller molecules can get through those pores much more quickly and end up much lower on the gel. Once the compounds are separated they are stained with a compound dye such as coomassie blue to visualize the results. 8). The key point of this question is what is SDS Page used for? It asks SDS Page would be least effective in distinguishing between the masses of different? Well the passage tells us it is good for proteins so we can cross off all the answer choices that our proteins which B D. This leaves us with only A. Why does A make sense? A states carbohydrate-rich glycoproteins. Does the passage say Sodium Dodecyl suflate can disrupt the covalent bonds in carbohydratesno! 9). Coomassie blue allows the results of electrophoresis to be visualized. 10. Which of the following proteins would move the most slowly through the gel in SDS page. Well we know from our explanation of the passage that larger molecules move more slowly through the gel because they cannot get through the pores. If you didn't know this it also said it in the passage that The rate of movement through the gel is inversely proportional to the logarithm of the molecular weight of the proteins 11. Key point here is that SDS is unnecessary in analyzing nucleic acids because nucleic acids already contain negatively charged phosphate groups in proportion to their size. 12. Given that SDS does not cleave covalent bonds, which protein structure cannot be disrupted by SDS? We know that primary structure involves covalent bonds, peptide bonds to be exact so our answer is A. Primary structure. 13. What would happen if a protein structure with quaternary structure or multiple subunits were exposed to SDS page. Well let's think SDS page disrupts non-covalent interactions. The mercatoethanol added would also rip apart disulfide bonds. What are the five forces that hold together quaternary structure? 1 Disulfide Bonds 2 Electrostatics 3 H-bond, 4 Van der Waals 5 Hydrophobic interactions. So if SDS and mercaptoethanol can disrupt these interactions then the subunits will be broken apart and travel through the gel separately according to their size. 14. So now that we have gone through this entire technique what is the limitation to SDS page in identifying different proteins with a protein mixture? SDS page only separates proteins by size. So what if we have different proteins that are of the same or similar size. The SDS page technique would not be able to separate them. Answer choice A is out-of-scope and simply does not make sense. How is it likely to be an expensive process that was never discussed in the passage. C is true but those proteins can also be spontaenously refolded and is not a limitation in identifying proteins. It may be a limitation in extracting proteins.

This passage is mainly talking about glycolysis which is a topic you should be an expert in. Let's describe the process of glycolysis: Glucose first enters the cell through facilitated diffusion down it's concentration gradient. Once inside the cell, it is phosphorylated by hexokinase to Glucose-6phosphate. Glucose 6-phosphate can then enter glycolysis or be converted to glycogen. If it enters the glycolysis, glucose-6-phosphate is isomerized into fructose-6-phosphate then fruc-6-phos- is converted to Fructose 1,6- bisphosphate which is the committing step in the process. Once this step occurs the body has to commit this molecule to glycolysis. These steps are the investment steps where two molecules of ATP are invested. Next fructose-1,6-bisphosphate is split into dihydroxyacetone and glyceraldehyde-3-phosphate (PGAL). Dihydroxyacetone is converted to PGAL. PGAL is shuttled into the next step of glycolysis when an oxidation reaction occurs where a phosphate group is picked up and NAD+ is reduced to NADH. Then that phosphate group that was just picked up is used to make ATP in a process called substrate level phosphorylation. Several other reactions occur and the remaining phosphate is used to make another molecule of ATP. Thus we have made 4 ATPS (2 per PGAL molecule) but our net production is 2 ATPs and 2 NADHs. 2 Pyruvate molecules are released.

Lecture 3 In-Class Exam Summary of Passage Disease causing agents are called Pathogens. Sterilization is the removal or destruction of all living cells, viable spores, viruses, and viroids. Sometimes it is only deemed necessary to kill or inhibit pathogens. This is called disinfection. Sanitation reduces the number of microbes to levels considered safe by public health standards. Obligatory anaerobe- Oxygen is poison to such organisms. They can only exist in an environment without oxygen- obligatory! Facultative anaerobes- Normally perform aerobic respiration with oxygen but can also switch to anaerobic conditions and survive without oxygen, using fermentation or other similar processes. Prime example- Yeast! 47 Certain eating utensils are treated with a sanitizer. After 3 minutes the number of microbes is reduced from 6.25 x 10^12 to 2.5 x 10^11. According to the passage 3 minutes more exposure to the sanitizer would reduce the number of microbes to... S1 If after 3 minutes the number of microbes reduced from 6.25 x 10^12 to 2.5 x 10^11then what would be the number of microbes left after another 3 minutes of treatment with sanitizer. Give: 6.25 x 10^12 to 2.5 x 10^11 we have a reduction rate where 96% of the bacteria are killed. It is not likely that you would have been able to calculate 96% exactly. S2 Let's look at the answers. From the calculation above we know that A. 1.0 x 10^10 1/25 = .04 or 4% this is our answer. B 5.0 x 10^10 C 6.0 x 10^10 D 1.9 x 10^11 D is way off. It is too close in value to 2.5 x 10^11. We are looking for a reduction by 96%. For this type of question, only quick and efficient calculations will save you. Always make sure that you understand what's being asked first before approaching the question. 48 From Table 1, the D value for Salmonella in Chicken a la King at 70 C is: S1 From table 1, the D value for Salmonella at 70 C is what? Looking at table 1 we see that we have to extrapolate using the z value. The D value at 60 C is 0.40. Z value is 5.0. So we know that the z value is the increase in temperature necessary to reduce a D value by a factor of 10. So by going to 70 we are reducing the D value by a factor of 100. So .40/100 = .0040 mins x 60 secs/mins - .24 secs. A. 0.24 secs- Thus our answer is A. I got this one wrong because I used the wrong category. Make sure you pay attention. B 2.4 secs C 24 secs D 40 minutes 49. If federal regulations require that canned food be heated at 121 C long enough to reduce a colony of C. botulinum in a phosphate buffer from 10^12 bacteria to 1 bacteria, how long must canned food be heated. 51 According to the passage, which of the following is true concerning a surface that has been disinfected? S1. According to the passage what happens when a surface is disinefected. Let's look back in the passage. Disinfection is on the left half in the middle. Sometimes it is only deemed necessary to kill or inhibit pathogens. This is called disinfection.

S2 Ok let's look at the answer choices. A No living microorganisms exist on the surface. - Wrong. It is sterilization where all living cells are killed. B Some living microbes remain on the surface, but all or most microbes capable of producing disease have been destroyed to reduced. - This makes sense and goes along with our evidence from the passage. This is the best answer! C The surface has been cleansed of bacteria, but not necessarily viruses or viroids. The passage states it kills or inhibits pathogens which includes viruses and viroids. This was explained in the first paragraph. Thus this answer is wrong. D All pathogens on the surface are destroyed or removed, while all nonpathogens on the surface remain alive. No distinction is made here about how all non-pathogens are spared in the process. 52 Which of the following statements is best supported by the data in Table 1? S1 Which of the following answer choices is best supported by data in Table 1? S2 let's look at the answer choice and look to see if they are supported in Table 1 A) A bacterium's resistance to heat is directly related to its z-value.- This is not true. If we look at all the values at 60 C we see that S. aureus on turkey stuffing which has a d value of 15 mins meaning it has a high heat resistane but its z-value is only 6.8. Salmonalla which has a d value of .40, low heat resistance but has a competitive z value of 5.0. Also there are using different subtrates so nothing can even be assumed because there is no control. B) C. botulinum is more likely to contaminate commercially processed food than S. aureus- OS just because the passage talks specifically about C. botulinum doesn't make this true. The passage never says this. This is Out-of-scope. C) A bacterium's resistance to heat may vary depending upon its environmentI feel as if this was stated in the passage. Environmental factors may affect D values And yes! It is supported by table 1. S aureus has different heat resistances for the different substrates this is our answer! D) In chicken a la king, Salmonella is more resistant to heat than S. aureus. We see that this answer choice is just plain wrong. 53 Which of the following statements does NOT contradict the information in the passage concerning C. botulinum? S1: Which of the following answer choices does not contradict the information in the passage about C botulinum or which of the answer choices are true according to the passage about C botulinum. Let's look at where this was stated which was in the last paragraph. C bot. Is an obligatory anaerobic, gram positive bacterium found in soil and aquatic sediments...it can be effectively treated with an injection of antibodies produced by horses A C. botulinum thrives in the presence or absence of oxygen. This is wrong or contradicts the passage. C bot is an obligatory anerobe meaning it needs the absence of oxygen. Oxygen is poison to it! B C. botulinum has a lipid bilayer outside its peptidoglycan wall.- This is also wrong and contradicts the passage. C. botulinum is a gram-positive bacteria and so it does not have a lipid bilayer outside its peptidoglycan wall. C A glass containing C. botulinum may be disinfected by immersion in boiling water moist heat! Directly supported by the passage. D Animals have no natural defense against neurotoxin produced by C. Botulium. This is not true because antibodies from horses are used to treat the disease.

54 One strand of an IS element begins with the nucleotide sequence 5'-ACTGTTAAG-3'. The same strand must end with the nucleotide sequence: S1: If one strand of an IS(insertion sequence) element begins with the nucleotide sequence 5'....., what must the other end be. The setup of the questions tells us that this was explained in the passage. Let's look at P2. Each single strand of DNA in an IS element possesses a nucleotide sequence at one end that is complementary to the reverse sequence of nucleotides at its other end Ok so we know first that is complementary so 3' TGACAATTC- 5' The reverse of this so that is in in the 5' to '3 direction is 5'-CTTAACAGT-3' S2 So we are looking for an answer choice that is the nucleotide sequence above Our answer is C! 55 When the passage states that transposons are similar to retroviruses, to what aspect of the life cycle of a retrovrius is the passage most likely referring? S1 According to the passage how are transposons similat to retroviruses? Which aspect of the life cycle of a retrovirus is the passage referring to? Key words are transposons are similar to retroviruses, what aspect of the life cycle? S2 Let's look back in the passage. They are similar to retroviruses How, well they insert themselves into the bacterial chromosome just as how prophages do. A. reverse transcription of viral RNA- Wrong. You don't reverse transcribe RNA. And, it is not stated that the the TGE's use reverse transcriptase. B Proliferation of multple copies of viral genome leading to the lyses of the host cell. This is lytic phase. Transposons don't lyse the cell. C. capsid formation. Transposons don't form capsids! They just stay within the genome. D. The procedure by which the viral genome integrates into the host gennome. Yes! And there is support for this in the passage. 56 Which of the following is not typically associated with genetic recombination in prokaryotes? S1 Which of the following answer choices is not typically associated with genetic recombination in prokaryotes. What are the three forms of genetic recombination we know of in bacteria? 1) Conjugation 2). Transformation 3) Transduction S2 Let's look at the answer choice. A Transduction- This is one of the methods we listed so this is not our ansewr. B TGE we just read about this in the passage. -reshuffling of genetic material. C binary fission- produces identical daughter cells in bacteria- This is our answer! D transformation- nope. That is a form of genetic recombination. 57 Which of the following mechanisms of genetic recombination between prokaryotes involves plasmids? S1 Which of the answer choices is a mechanism of genetic recombination between prokaryotes that involves plasmids? This is almost a stand-alone question. Remember to focus on key words for standalone category questions. Which method do we know involves plasmids? Conjugation! A Transduction- This is the process of genetic recombination through viruses. Virus mediated genetic recombination. B Conjugation- Yes, this is our answer! C Meiosis- Prokaryotes don't perform meiosis or mitosis.. D Binary fission- This produces identical daughter cells. Remember binary fission is the mitosis of Prokaryotes!

58 Bacterium A is able to live on a histidine deficient medium. Bacterium B is not. After initiating conjugation with Bacterium B, Bacterium A is unable to live on a histidine deficient medium. Which of the following statements is most likely true concerning the conjugation? S1. Which of thefollowing answer choices probably occurred from the conjugation or which is most likely true? So Bacterium A performs conjugation with Bacterium B and then loses the ability to make histidine. Perhaps Bac. A lost its plasmid. S2 I wasn't paying attention and don't know what conservative transposition or replicative transpostions is. Well I know I can look back in the passage let's look for it. The last paragraph. A TGE may move as a complete entity, called conservative transposition, or it may move a duplicate copy of itself, called replicative transposition So we know what we are looking for is conservative transposition because Bac A lost its plasmid or DNA in this case. A Conservative Transposition.. A is our answer. 59. While integrated into the host cell DNA a lamba phage is called a: S1: Clear. A phage integrated into the chromosome is called a prophage similar to a provirus S2. Let's look at the answer choices. A- Wrong. Virion is used interchangeably to describe a virus. B- Prophage. Bingo! C Chromosome- NS D- Plasmid- What. I guess this would be a deceptive answer choice if you didn't know the difference between a plasmid and a phage. 60. Lamba phage most likely integrates into the host cell DNA in the: S1 Where does the lamba phage integrate into the host cell DNA? Since it is a phage, in the nucleoid (chromosome of the bacterium) S2 Let's look at the answer choices A- host cell nucleus- Wrong. Bacteria don't have nuclei B- host cell mitochondria- Wrong. Bacteria don't have mitochondria either. C- lumen of the endoplasmic reticulum -NS. Bacteria don't have complex, membrane bound organelles and why would it be in the lumen of the ER. Is there DNA there? D. Cytoplasm of the host cell. Yes, this is where the bacterial chromosome or nucleoid is. 61. Which of the following would most likely lead to lysis of a host cell infected with lamba phage in the lyosgenic stage? S1.clear enough. Let's look in the passage. Damaged DNA activates a cellular protease that degrades lamba repressor. When the concentration of lamba repressor falls below the critical limit... So we are looking for something causes DNA damage. A-Infection by a second lambda phage. This doesn't really cause DNA damage, just genetic recombination. B-binary fission. This would produced identical daughter cells- NS C-Exposure to Ultraviolet radiation.- Yes, this makes sense. UV radiation causes Thymine dimers to form which damages DNA D- Mitosis- Wrong. Bacteria do not perform mitosis. 62. Which of the following must be true in order for infection with a lambda phage to take place? S1: What needs to happen before the phage can infect a bacterium? This is pretty much a stand-alone question. We know that the phage must first adsborb onto the cell wall of the bacteria by binding to a specific receptor.

B. Bingo! 63.