Question

3. Explain the process of DNA replication, including how it begins and the reasons for the directionality of the process (5’ to 3’). What are the major enzymes involved in the process, and what individual functions do they have that allows for replication to occur?

a) What are the major differences between prokaryotic and eukaryotic DNA (what is different about how the DNA is organized)?

b) What kinds of mistakes can happen during replication or types of damage that can occur to DNA (types of mutations)? How can these mistakes be repaired?

Answer 1

Ans 3:- The DNA replication occurs in three steps which are as:-

DNA REPLICATION IN PROKARYOTES

(i) Initiation (ii) Elongation (iii)Termination

Initiation:- DnaA protein intitates replication in E.Coli at oriC. DnaA binds the 9-mers sequence in oriC, forms the initial complex, and facilitates the initial strand seperation, or melting, of E.Coli DNA, which occurs at 13-mers and thus forms the open complex. Further melting of thw two strands of the E.Coli chromosome to generate unpaired template strands is mediated by DnaB proteins, a helicase and clamps around the single strands of the DNA in open complex now protein DnaC comes and acts as the helicase loader and hence seperates the strand further with the opening of the strand the single strand binding protein gets attached to the single strands and hence prevents the further annealing of the strands. DNA polymerses can only elongate existing primer strands of DNA or RNA. the primers used during DNA replication are synthesized by the RNA polymerase primase . After the bound primases synthesize short primer RNAs complementary to both strands of duplex DNA they dissociate from the single stranded template. Sequence during initiation is as:

1. Binding of DnaA protein to oriC

2. Loading of DNA helicase

3. helicase open helix and binds primase to form primosome

4. Synthesis of RNA primer

5.Initiation of DNA polymerization by DNA polymerase.

Elongation:- DNA polymerse catalyse the addition of deoxyribonucleotide units to DNA chain.the chain elongation reaction catalyzed by the DNA pol. is a nucleophilic attack by the 3'-hydroxyl group of the primer on the inner most phosphorous atom of the deoxyribonucleoside triphosphte. A phosphodiester bridge forms with the concomitanr release of pyrophosphate. the catalytic megnesium metal ion in the active site plays an importan role during the elongation process.the elongation of the DNA chain proceeds in the 5'-3' direction. At each growinf fork, one strand, called the leading strand is synthesized continuously from a singlr primer on the leading strand template and grown in the 5'-3' direction. synthesis of the lagging strand is more complicated becaose DNA pol can only synthesixe in 5'-3' only adding nucleotide only to 3' end of a primer or growing DNA strand. movement of growing fork unveils the template strand for lagging strand synthesis in the 5'-3' direction. short peices of DNA called okazaki fragments are formed. the RNA fragments of the previous is removed by the 5'-3' exonuclese activity of DNA pol. I. This enzyme also fills the gaps between the lagging strand fragments which then are ligated together by DNA ligase.

Termination:- the termination occurs at the ter sequences which are 22bp long sequences called the ter sequences in E.coli has TerA-TerF. The two replicating forks approach eachother as their occurs a bidiretional replication in the ecoli and tangling of the two newly synthesized circular DNA occurs which are relieved by the Topoisomerase IV and thus termination ends.

Ans (a) in prokaryotes the genome is concentrated in the nucleoid while as eukaryotes in nucleus and mitochondria.

(b) prokaryotes only contains genes(exons) eukaryotes contain both introns and exons.

(c) prokaryotes conation circular DNA eukaryotes have linear.

(d) extrachromosomal plasmids are not present in eularyor=tes.

(e) transcription and translation occurs simultaneously in prokaryotes while as there are diffrent compalrtment in the eukaryotes for transcription and translation.