7/29/2019 330ex110

1/4

Name___________________________ UT EID_________________________

BIO 330Exam 1

February 23, 2010

Read each question carefully before answering. Many questions have multiple parts.

1. Name at least two cellular proteins that are required prior to the initiation of poliovirus RNAsynthesis. (5 points)

Why do some picornaviruses require adsorptive endocytosis for penetration and uncoatingwhereas other members of this same family do not? (5 points)

2. What is a negative polarity viral RNA? (4 points) Explain why this RNA would not beinfectious. (4 points)

3. A new virus (ACHOO2) has been identified from the nasal washings of children with cough,sneezing, and fever. You determine that the ACHOO2 virus has an icosahedral capsid withT=1. How did you do this? (4 points)

Draw one of the triangular faces and show the 5-fold, 3-fold, and 2-fold axes of rotationalsymmetry. Also show positions of pentons and hexons. (6 points)

Are the interactions between subunits in this virus equivalent or quasi-equivalent? Why? (5points)

1

7/29/2019 330ex110

2/4

4. Since you appear to have a new virus, you would like to know the receptor used by ACHOO2.You attempt to identify the receptor using receptor competition experiments. Using a humanlung cell line, you first use a large amount of a known non-radioactive virus to bind to the cellsunder conditions where the virus cannot penetrate the cells. Once the first virus has bound, youthen add a second radioactive virus to determine binding. All unbound virus is removed fromthe cells by washing with non-radioactive medium. Subsequently, you determine the amount of

radioactive virus bound. You obtain the following results.Amount of radioactive virus bound (counts per minute)

Non-radioactivevirus used forreceptorsaturation

ACHOO2 Poliovirustype 1

HRV-2 HAV HPEV1

ACHOO2 230 10,401 345 11,455 12,345

Polio type 1 13,004 329 11,589 11,867 11,423

HRV-2 312 10,776 344 13,835 12,342

HAV 12,435 12,953 10,899 267 12,376

HPEV1 10,357 11,321 12,786 13,824 421How many receptor groups are represented in this experiment? Give your rationale. (6 points)

You use the same radioactive virus to determine intermediates in virus uncoating. The virus(150S) has been radiolabeled with 14C-amino acids and 3H-uridine. You incubate the virus withsusceptible cells for 30 minutes under conditions in which you prevent virus replication. Thenyou recover three sets of particles from the cells and analyze them on sucrose gradients after

ultracentrifugation and fractionation of the gradients. You observe: 140S particles that have both14C and 3H radioactivity, 120S particles that have both 14C and 3H radioactivity, and 90S particlesthat have only 14C radioactivity. Give a brief description of the order of events. (6 points)

5. What is the effect of the drug pactomycin on mammalian cells? (5 points)

What would be the purpose of using pactamycin mapping for the ACHOO virus? (5 points)

2

7/29/2019 330ex110

3/4

6. You decide to determine the concentration of ACHOO virus in nasal washings of youngchildren. Subsequently, you incubated the ACHOO virus in the presence of pH 3.0 buffer andin the presence of pH 7.4 buffer, and then you diluted the virus and 0.2 ml of each dilution wasincubated at 37oC with human lung fibroblasts. After incubation for 1 hour with the virusdilutions, media is added and the cells were further incubated at 37oC for 48 hours.

You observe the following results.Number of plaques

Dilution pH 3.0 buffer pH 7.4 buffer

1:1000 2 Too many to count1:10,000 1 389

1:100,000 0 43

What is the average titer of the virus? Show your calculations. (5 points)

What does the appearance of plaques reveal about the replication of ACHOO virus? (5 points)

What did the addition of pH 3 buffer tell you about the likely method of human infection byACHOO virus? (5 points)

7. Why would you consider a virus to be a living organism? (6 points)

What are two basic components that EVERY virus has? (4 points)

3

7/29/2019 330ex110

4/4

8. Name two characteristics that differ between cell lines and transformed cell lines. (4 points)

Why is telomerase active in both types of cells? (4 points)

9. Since poliovirus RNA lacks a 7-methyl guanosine (cap) at the 5 end, how can the RNA betranslated? Be specific. (6 points)

How does poliovirus inhibit host cell translation? Be specific. (6 points) Graduate students: Inyour answer, explain a possible mechanism where cellular mRNAs can escape translationinhibition in poliovirus-infected cells.

4