Q.25 The enzyme which adds phosphate group to the free 5′ terminus of a DNA
sequence is
(A) adenosine kinase
(B) alkaline phosphatase
(C) polynucleotide kinase
(D) terminal deoxynucleotidyl transferase
Polynucleotide kinase is the correct enzyme for adding a phosphate group to the free 5′ terminus of a DNA sequence. This multiple-choice question tests knowledge of key enzymes in molecular biology, particularly those involved in DNA end modification.
Correct Answer
(C) Polynucleotide kinase
Polynucleotide kinase (PNK), often exemplified by T4 PNK from bacteriophage T4, catalyzes the transfer of the gamma-phosphate from ATP to the 5′-hydroxyl (5′-OH) group of DNA or RNA, generating a 5′-monophosphate end essential for ligation reactions. This activity is crucial in DNA repair, cloning, and labeling procedures, where unphosphorylated ends must be activated.
Option Breakdown
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(A) Adenosine kinase: This enzyme phosphorylates adenosine to AMP using ATP, playing a role in nucleoside salvage pathways but not in adding phosphates to DNA termini.
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(B) Alkaline phosphatase: It removes phosphate groups from the 5′ end of DNA or RNA (dephosphorylation), the opposite of the required function, and is used to prevent self-ligation in cloning.
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(C) Polynucleotide kinase: Correct, as its 5′-kinase domain specifically phosphorylates free 5′-OH ends on nucleic acids.
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(D) Terminal deoxynucleotidyl transferase: This template-independent polymerase adds deoxynucleotides to the 3′ hydroxyl end of DNA, used for tailing but not 5′ phosphorylation.
