5-Bromovinyl 2'-deoxyuridine phosphorylation by mitochondrial and cytosolic thymidine kinase (TK2 and TK1) and its use in selective measurement of TK2 activity in crude extracts

Nucleosides Nucleotides Nucleic Acids. 2008 Jun;27(6):858-62. doi: 10.1080/15257770802146510.

Abstract

Mitochondrial thymidine kinase (TK2) is responsible for phosphorylation of thymidine and deoxycytidine and plays a crucial role in mitochondrial DNA precursor synthesis. TK2 is expressed in all tissues at low levels complicating accurate determinations, especially in tissues with high cytosolic thymidine kinase (TK1) activity. Recently, 5-bromovinyl 2 '-deoxyuridine (BvdU) at 0.2 micro M was used to measure TK2 activity selectively. BvdU phosphorylation by pure human TK2 and TK1 was tested here, and the ratio of BvdU phosphorylation by TK2/TK1 was 91 at 0.2 micro M but was 500 at 2.5 micro M. Therefore, for reliable measurement of TK2 activity higher BvdU concentration should be used.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bromodeoxyuridine / analogs & derivatives*
  • Bromodeoxyuridine / metabolism
  • Complex Mixtures / metabolism*
  • Cytosol / enzymology*
  • Enzyme Inhibitors / pharmacology
  • Humans
  • Mice
  • Mitochondria / enzymology*
  • Organ Specificity
  • Phosphorylation
  • Substrate Specificity
  • Thymidine / metabolism
  • Thymidine Kinase / metabolism*
  • Thymidine Phosphorylase / antagonists & inhibitors

Substances

  • Complex Mixtures
  • Enzyme Inhibitors
  • brivudine
  • Thymidine Phosphorylase
  • thymidine kinase 2
  • Thymidine Kinase
  • thymidine kinase 1
  • Bromodeoxyuridine
  • Thymidine