What is exonuclease activity?
Terminology: The ability to remove nucleotides one at a time from the end of a chain is called exonuclease activity. (exo = from the exterior or end). There are two types of exonuclease: a. The enzymatic ability of DNA polymerase used in proof reading removes nucleotides one at a time from the 3′ end of a chain.
What is the purpose of an enzyme’s 3/5 exonuclease activity?
The 3’–5′ exonuclease activity of the enzyme allows the incorrect base pair to be excised (this activity is known as proofreading). Following base excision, the polymerase can re-insert the correct base and replication can continue forwards.
Which of the following enzymes has 5 to 3 exonuclease activity?
DNA polymerase I is a single polypeptide chain with 928 amino acids and molecular weight of 109 kDa. It has three sites, which provide three distinct catalytic activities: 3′ to 5′ exonuclease, 5′ to 3′ exonuclease, and 5′ to 3′ polymerase.
What is the purpose of the 5 to 3 exonuclease activity of DNA polymerase I?
DNA Polymerase I possesses a 3´→5´ exonuclease activity or “proofreading” function, which lowers the error rate during DNA replication, and also contains a 5´→3´ exonuclease activity, which enables the enzyme to replace nucleotides in the growing strand of DNA by nick translation.
What activity has 3 to 5 exonuclease?
The 3′→5′ exonuclease activity intrinsic to several DNA polymerases plays a primary role in genetic stability; it acts as a first line of defense in correcting DNA polymerase errors. A mismatched basepair at the primer terminus is the preferred substrate for the exonuclease activity over a correct basepair.
What would happen if a mutation eliminated the 5 3 exonuclease activity of DNA polymerase I?
What would be the effect on DNA replication of a mutation that destroyed the 5’3′ polymerase activity in DNA polymerase I? There would be a loss of the ability to synthesize DNA in the gaps created by the removal of RNA primers.
What is the specific activity of Taq polymerase?
One such form, the Stoffel fragment, was constructed by deleting the first 867 bp of the Taq DNA polymerase gene, to yield a 544– amino acid protein with an inferred molecular weight of 61,300 and a specific activity of 369,000 U/mg ( Lawyer et al., 1993; see also Table 1 ).
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What kind of activity does pfu polymerase have?
Pfu is a polymerase with 3′-5′ exonuclease activity meaning that as the PCR product is synthesized it checks which nucleotides have been incorporated and can work backwards to remove incorrect bases and replace them with the correct ones.
When does polymerase activity occur in a PCR reaction?
This low-temperature polymerase activity is the basis for nonspecific amplification associated with mispriming events that occur during the initial phase of the PCR reaction. Extension can occur from oligodeoxynucleotide primers that anneal nonspecifically to template DNA before the first denaturation step at 93–95°C.