Protein Modeling · Division C · GGSO Invitational · 2021
75 questions · 211.5 points · 50 min· 12 questions from the original omitted
Original test sourceRead each question carefully. Some of the question stems provide specific instructions on how to input your answers. This is because much of the test (multiple choice/answer and fill in the blank sections) will be autograded and we want to make sure you're getting full credit for all of your correct answers! :) Note that the fill-in-the-blank autograder is NOT case sensitive.
Please try to be concise! All of the short answer questions are able to be answered in less than 3-4 sentences. Most of these questions specify a limit and unfortunately, we will not be giving any credit to answers that surpass this limit.
The grading scheme is as follows: 50% Computer exploration, 50% exam. The computer exploration section is at the end of the exam.
Have fun!!!
You have discovered a mutation of protein that normally gets phosphorylated at a certain tyrosine residue but has been mutated to a glycine. What amino acid mutation(s) at the same residue would best rescue the wild type phenotype?
Select all mutations that would most likely lead to a loss of overall protein function:
The next 6 questions describe different biological processes. For each, determine whether the driving force of the process is due to A. hydrogen bonding, B. the hydrophobic effect, or C. covalent bonding. Some of the answers may be used more than once or not at all. Please only write the letter corresponding to the answer choice ("A" for hydrogen bonding, "B" for hydrophobic effect, or "C" for covalent bonding).
Nsp1 recognizes and binds to host RNA
Formation of vesicles in the cell
Ubiquitination of cdc20
DNA tertiary structure
(For the next 4 questions) You run a protein on an SDS-PAGE gel without any reducing agent and find one band of ~150 kDa. When you run the same protein on an SDS-PAGE gel with a reducing agent, you find two bands, one at ~50 kDa and one at ~25 kDa.
What can you deduce about the function of this protein or where it acts? (1 sentence answer)
Sequencing of the gene for this protein reveals hotspots of mutations as a result of cytidine deaminations. What protein is this?
You also generate a cDNA from the mRNA transcript of this protein. Do you expect the cDNA to be longer, shorter, or about the same length as the gene? Why?
You then sequence the same gene in a cell from the same organism that does NOT normally express this protein. You find that the gene is much longer in this cell than in the cell that expresses this protein. Explain this phenomenon in 3 sentences or less. (hint: this is related to the identity of the protein)
(For the next 2 questions) Below are two sample Ramachandran plots for reference. A shows the allowed conformation angles of different secondary structures while B shows the Ramachandran plot for the general amino acid.
Shown is the Ramachandran plot for glycine. Explain in three sentences or less why the glycine plot looks different from the plot for most amino acids.
Shown is the Ramachandran plot for proline. Explain in three sentences or less why the proline plot looks different from the plot for most amino acids.
Select all of the following statements that are true about protein folding.
Resonance causes peptide bonds in proteins to become
Rotation around which of the following bonds are permitted?
Most of the aromatic residues are found in
Select all of the following polypeptides which will have a neutral charge under physiological conditions
Which of the following kinds of beta sheets are the most stable.
A protein is placed in an environment that is 95 degrees celsius. Which of the following parts of the protein structure are likely to be affected?
(Next two questions) The CFTR protein is a transmembrane protein that acts as a chloride channel in various cells of the body.
Knowing this, what is one structural feature that you expect to find?
What other information does the peptide need to contain to ensure proper localization in the cell?
Researchers wish to synthesize a protein in vitro. After translating this protein, the protein takes many days to fold. However, in vivo this protein folds very quickly. Select all of the following reasons that could cause this difference.
In order for alpha-helices to form
London dispersion forces contribute to protein formation by
Which domain(s) of Cas9 are mutated in dCas9?
Why are uracil glycosylase inhibitor proteins often included in cytosine base editors?
Cas9 nickases usually only have one of the two mutations seen in dCas9. For the next 2 questions, state which mutation and in what domain you should use if you want to cut the:
WRITE YOUR ANSWERS AS: Mutation in the LEFT BLANK, the domain in the RIGHT BLANK. The mutation should follow the proper notation shown below (i.e. what's shown in papers).
Non-target strand: Mutation ____, Domain ____
Target strand: Mutation ____, Domain ____
Fluorescent in situ hybridization (FISH) uses a fluorescently labelled probe that binds to a complementary sequence of nucleic acid to visualize specific genomic loci. Fluorescence microscopy can be used to localize the bound probes, allowing for researchers to examine spatial-temporal patterns of gene expression. You, an astute Protein Modeling competitor, realize that you could modify the CRISPR/Cas9 system to do something similar. Select everything you will need for this new FISH-like visualization technique.
For the next eight questions, determine whether an A. cytosine base editor, B. adenine base editor, or C. neither could be used to fix the defect. Some of the answers may be used more than once or not at all. Please only write the letter corresponding to the answer choice ("A" for cytosine base editor, "B" for adenine base editor, or "C" for neither).
Nota bene on mutation notation used below:
GENE c. 14 A>G
GENE c. 87 C>T
GENE c. 3589 G>C
GENE c. 24 T>C
GENE c. 77-78 ins T
GENE c. 80-83 dup
GENE c. *19 C>A [the * here means that this is after the stop codon]
GENE c. *3 G>A
For the next 4 questions: In the ear, cochlear hair cells detect sound through mechanical movement. Mechanosensory ion channels, such as that encoded by the transmembrane channel-like 1 gene (Tcm1), are crucial in relaying this mechanical movement into a signal that is able to be processed by neurons.
You sequence mice that have hearing loss and find a mutation in Tcm1. Based on the sequencing results below, what is the mutation? (Hint: look at WT (wild type) vs. mutant 2)
Can you explain the sequencing results for mutant 1 (one sentence or less)?
Your colleague wants to use CRISPR gene editing with a homology-directed repair approach to correct this defect. Would this work? Explain your answer in one sentence.
How would you correct this defect? Explain why your method would work. Answers will only get full credit if they are less than 3 sentences long and mention a specific enzyme.
In the bacterial immune system, the CRISPR-mediated defense
Select all of the following steps of CRISPR immunity that involves spacer acquisition
Select all of the following that are true about Type I CRISPR-Cas9 systems
In one sentence, explain how the bacterial immune system avoids auto-immune targeting of the CRISPR array.
Use these two figures to answer the following 5 questions.
Figure 1)
Figure 2)
The images investigate whether CRISPR spacer content determines phage resistance. The phage sensitivity of each S. thermophilus strain to phages 858 and 2972 is represented at the bottom of each figure as a bar chart of the efficiency of plaquing, which is the plaque count ratio of a mutant strain to that of the wild-type. Figure 2 shows nine different mutants that are resistant to phage 858 and/or phage 2972. Figure 1 shows two of these mutants that have further alterations to their CRISPR1 locus. These changes are as follows:
I, WT Φ 858 +S1S2 Φ 858 +S1S2 II, WT ΔCRISPR1 where CRISPR1 is deleted III, WT Φ 858 ::pR where a repeat was inserted between the cas genes and CRISPR +S1S2 IV, WT Φ 2972 ::pS1S2 where spacer S1 and S2 are added +S4 V, WT Φ 858 ::pcas5– where cas5 is inactivated +S1S2 +S1S2 VI, WT Φ 858 ::pcas7– where cas7 is inactivated
Explain what evidence in the figures supports the idea that proper spacers and cas genes alone do not provide resistance.
Next three questions. Tiebreaker: The following two figures showcase an arms race between a phage and a bacteria with CRISPR-based immunity of increasing resistance and increasing infectivity. In a broad sense, what do these two figures suggest the mechanisms are for each. A detailed understanding of the two figures is not necessary. For each figure in one sentence describe the mechanism and whether that figure refers to increasing resistance or infectivity.
Figure 1)
Figure 2)
Figure 1
Figure 2
Together, how do these two mechanisms impact the relationship between phage and bacteria?
You need to make bacteriophage (a virus that infects bacteria)-resistant mutants (BIM) of a bacterial strain you're working with that you know uses the CRISPR-Cas system for adaptive immunity. To check if your mutants have acquired spacers, you use PCR (polymerase chain reaction) to amplify the CRISPR repeat-spacer arrays of your mutants. You run your results on an agarose gel against a DNA ladder and the PCR amplicon of the CRISPR array of wild-type (WT), non-mutated strain. Explain in one sentence how you can tell whether or not your mutants have incorporated the spacers.
Name another way/experiment you could do to verify if the spacer has been added.
Use the jmol design environment to load each structure by the ID given: https://cbm.msoe.edu/markMyweb/jmolDesignEnvironment/#forward
(For the next 3 questions) Load 1T02 into Jmol. This enzyme is HMG-CoA Reductase, which is involved in the rate limiting step of cholesterol synthesis. Statin, a drug often used to lower cholesterol levels, is also shown interacting with the enzyme.
Which is the structure of statin?
The following residues all are involved in interacting with statin. For each, write the amino acid at that residue and predict its interaction with statin (a few words). You can refer to the specific residue by the corresponding highlighted letter.
66a267, chain A
66b85, chain A
66c261, chain A
(For the next 5 questions) Now, load 5N2F into Jmol. This shows PD-L1 bound to a small molecule inhibitor. PD-L1 is expressed on tissues and binds to PD-1 on cytotoxic T-cells to dampen their activity. PD-1/PD-L1 inhibitors are actively being used and studied for immunotherapy against certain cancers.
What kind of oligomeric protein is shown?
Take a look at the small molecule. How many heteroatoms are there and what are they?
Look more closely at the ring structure in the small molecule with the green atoms (green using the cpk color scheme). This ring has a special interaction with a certain residue. Name the chain, the amino acid, and the location of the residue.
(Continued from above) Tiebreaker: What is the name of this interaction?
What would happen if this protein were put into a reducing environment? Would there be any additional cysteine residues exposed and if so, how many?
Now load 4C9D to answer the next 8 questions. This shows Cas6 bound to a CRISPR repeat.
What is the residue number of the lysine that interacts with G28 of the RNA?
The loop containing the above lysine is rich in what amino acid (three letter code)?
What other amino acid on this loop interacts with the RNA (amino acid and number)?
The RNA forms what base pairing pattern?
How many nucleotide-nucleotide pairings are formed in the RNA structure questioned about above? The following two questions refer to the pairings described here
What do you notice about the aromatic rings of these bases in the nucleotide-nucleotide pairings of the above question?
In total, how many hydrogen bonds exist between the pairs of bases in the above nucleotide-nucleotide pairings?
One reason why it is important that Cas6B is a dimer has to do with the interaction with G17. What do you see that demonstrates this (be specific)?
Now, please load 5F9R to answer the last questions. This shows Cas9 in complex with an sgRNA and double stranded DNA.
Do the Cas9 residues Phe-916 and Leu-921 interact with the target or nontarget DNA strand?
At approximately what angle is the DNA helix bent?
The PAM occurs between bases 15 and 25.
First blank: What is the PAM sequence (5' to 3')? Write ONLY the nucleotides.
The PAM occurs between bases 15 and 25.
Second blank: What is the first base pair position of the PAM sequence?
The PAM occurs between bases 15 and 25.
Third blank: What is the last base pair position (higher number than second blank) of the PAM sequence?
How many bases of the RNA bind the DNA?
The largest beta sheet has how many strands?