Question

1.Both Poly A site choice and alternative splicing are used to produce two different hormones Calcitonin and CGRP. What is this Poly A directing site? State the mechanism of addition of the poly A tail.

A.

5’-AAACGTGC-3’; the transcript is cleaved 10-30 bases downstream of the poly A site and then a polyadenylate polymerase adds a poly A tail

B.

5’-AAACGTGC-3’; the transcript is cleaved 10-30 bases downstream of the poly A site and then the enzyme telomerase adds a poly A tail

C.

5’-UAAUUU-3’; the transcript is cleaved 100 bases downstream of the poly A site and then the enzyme polynucleotide kinase adds a poly A tail

D.

5’-AAUAAA-3’; the transcript is cleaved 10-30 bases downstream of the poly A site and then the enzyme polynucleotide kinase adds a poly A tail

E.

5’-AAUAAA-3’; the transcript is cleaved 10-30 bases downstream of the poly A site and then a polyadenylate polymerase adds a poly A tail

2. At what stage is the 5’ cap added to the newly-made primary transcripts? (Before or after splicing? Before or after the formation of the poly A tail?). Draw the structure of the 5’-cap in your scrapbook and state what bonds link the cap to the rest of the RNA.

A.

The 5’ cap is formed after splicing, but before the poly A tail formation; a 5’5’ tri-phosphodiester bond

B.

The 5’ cap is formed before splicing and before the poly A tail formation; a 5’5’ tri-phosphodiester bond

C.

The 5’ cap is formed before splicing and before the poly A tail formation; a 5’5’ monophosphodiester bond

D.

The 5’ cap is formed before splicing and before the poly A tail formation; a 5’3’ monophosphoester bond

E.

The 5’ cap is formed after splicing and after poly A tail formation; a 5’5’ tri-phosphodiester bond

3. Draw the structures of AZT and DDI in your scrapbook and identify the structural feature in each that enables it to work in slowing down the multiplication of the AIDS virus?

A.

Containing an azide group at 3’-position, AZT generates a nitrene that covalently links to and inactivates the AIDS virus reverse transcriptase, but DDI, which contains a hydrogen atom at the 3’ position binds non-covalently to and inactivates the reverse transcriptase

B.

Containing and azido instead of a hydroxy group at the 3’ position, AZT and DDI cannot extend the DNA chain through a 3’5’ phosphodiester bond. AZT binds to AIDS virus reverse transcriptase more strongly than dNTPs

C.

Containing an azido instead of a hydroxy group at the 2’ position, AZT and DDI cannot extend the DNA chain through a 2’3’ phosphodiester bond. AZT binds to AIDS virus reverse transcriptase more strongly than dNTPs

D.

Both AZT and DDI share structural features that stimulate autophagy in the host cells that causes degradation of the AIDS virus

E.

As nucleotide mimetics, they stimulate the host cell RNA polymerase to produce specific gene products that function to expel the AIDS viral DNA before formation of the packaged virus

Answer 1

1.Both Poly A site choice and alternative splicing are used to produce two different hormones Calcitonin and CGRP. What is this Poly A directing site? State the mechanism of addition of the poly A tail.

Answer: (E)

5’-AAUAAA-3’; the transcript is cleaved 10-30 bases downstream of the poly A site and then a polyadenylate polymerase adds a poly A tail

Mechanism of addition of the poly A tail

Capping & splicing are both RNA poly mediated while tailing is RNA transcript mediated process. There are some factors crucial for this process; Cyt F (Cleavage stimulatory factor), CpsF (Cleavage & polyadenylation specificity factor). This process occurs in multistep manner.

Step1 Extention of the RNA transcript (by Pol II) beyond the polyA tail addition site

Step 2 Cleavage of RNA transcript (at GU rich region by an endonuclease) at addition site of polyA tail and formation of new 3' end ie CA where PAP polymerase does polyadenylation. (mechanism Image attached)

Q 2. At what stage is the 5’ cap added to the newly-made primary transcripts? (Before or after splicing? Before or after the formation of the poly A tail?). Draw the structure of the 5’-cap in your scrapbook and state what bonds link the cap to the rest of the RNA.

Answer: (B)

The 5’ cap is formed before splicing and before the poly A tail formation; a 5’,5’ tri-phosphodiester bond

The structure of the 5’-cap and bonds linking the cap to the rest of the RNA.

Capping is present ar 5' end of RNA and is attached with the 5',5'- triphosphate bond (between m7 of G residue & 5' end of RNA).

Q 3. Draw the structures of AZT and DDI in your scrapbook and identify the structural feature in each that enables it to work in slowing down the multiplication of the AIDS virus?

Answer: (B)

Containing and azido instead of a hydroxy group at the 3’ position, AZT and DDI cannot extend the DNA chain through a 3’,5’ phosphodiester bond. AZT binds to AIDS virus reverse transcriptase more strongly than dNTPs

Structure of

a) 3′-azido-3′-deoxythymidine (AZT)

b) 2′,3-dideoxyinosine (DDI)​​​​​​​