Fig. 2-4.2.1: Conversion of 5' dephospho-(deoxy) ribonucleic acid to 5' phospho-(deoxy) ribonucleic acid by the action of PNK.

The efficiency of phosphorylation is less in exchange reaction compared to forward reaction. Along with the phosphorylating activity, PNK also has 3' phosphatase activity.

There are two major uses of PNK:

• The linkers and adopters are phosphorylated along with the fragments of DNA before ligation, which requires a 5' phosphate. This includes products of polymerase chain reaction, which are generated by using non-phosphorylated primers.
  
  
• PNK is also used for radio labelling oligonucleotides, generally with ³²P for preparing hybridization probes.

PNK is inhibited by ammonium ions, so ammonium acetate cannot be used to precipitate nucleic acids before phosphorylation. Sometimes phosphate ions or NaCl of greater than 50 mM concentration can also inhibit this enzyme.