Question

3) a) Arguably, the way prokaryotic operons are structured has evolutionary advantages. Based on our discussions in class, briefly describe one advantage and one disadvantage of gene expression regulation in prokaryotes. Provide appropriate examples to strengthen your argument (3 pts).

b) Restriction enzymes, which are extensively used in molecular biology, are products of bacterial “immune” system. Since bacterial genomes span several million base pairs (E. coli > 4 million bps), and the presence of restriction site within a genome is more than likely, how does a bacteria manage to protect itself from innate REs? (1 pt )

3 d) Assuming the equal distribution of 4 nucleotides within a genome and a RE palindrome having C:G = A:T 6-base recognition sequence, what is the likely size of the fragment of the DNA treated with this particular RE? (2 pts)

E) What is the difference between type I and type II restriction enzymes, and how would you utilize each in a molecular biology experiment? Provide a simple hypothetical example for each. (3 pts)

Answer 1

3b. We know that in the prokaryotes an important enzyme known as the DNA methytrasferase is present. This is a very fast acting enzyme so when ever the new DNA strand is produced it methylates the newly formmed DNA this methylation process helps the prokaryotes by protecting their genome from their own restriction endonucleases that is present in their cytosol. The RE enzymes are quite slow enzymes mainly they are one of the slowest enzymes and requires time for their cutting in the sites so after the newly formed DNA the RE enzymes gets very little time to work on them or no time at all as the newly formed DNA is methylated at very fast rate.

E. The difference between type I and II RE enzymes are-

1.The type I RE cleaves the DNA at a site of 1000bp away from the restriction site whereas the type 2 cuts specifically at the restriction site.

2.The type I use ATP and ADoMET as the cofactors for their cutting but the type II do not require any such co factors and they only require Mg⁺⁺ as their cofators.

3. The type I functions both as a methylase and a restriction enzyme but the type II has no methylase activity.

In the case of mol biology labs only type 2 RE is used because it specifically cuts at its recognition site and produces either a blunt or a sticky end so they are used for cutting the long genomic DNA into smaller fragments before running them in gel electrophoresis or they are also used to insert and clone DNA fragments in to the artificial plasmids.

Type I is used when we need to digest a DNA fragment at random places or far away from the recognition sites.