Ly. Wang et al., SUBSTRATE-SPECIFICITY OF MITOCHONDRIAL 2'-DEOXYGUANOSINE KINASE - EFFICIENT PHOSPHORYLATION OF 2-CHLORODEOXYADENOSINE, The Journal of biological chemistry, 268(30), 1993, pp. 22847-22852
Mitochondrial deoxyguanosine kinase (dGK) (EC 2.7.1.113) was purified
to apparent homogeneity from bovine brain. The molecular mass of the n
ative protein was 56 kDa, as judged by gel filtration, and one single
band of 28 kDa was seen in sodium dodecyl sulfate-gel electrophoresis.
2'-Deoxyguanosine (dGuo) (K(m), 7.6 muM), 2'-deoxyinosine, and 2'-deo
xyadenosine (K(m), 60 muM) were substrates for the enzyme as well as s
everal dGuo analogs containing a lipophilic substituent at C-2'. Carbo
cyclic dGuo, 9-beta-D-arabinofuranosylguanine, 9-beta-D-arabinofuranos
ylhypoxanthine, and 9-beta-D-arabinofuranosyladenine were substrates f
or the enzyme, whereas no 3'-modified dGuo analogs were effective. Int
erestingly, 2-chloro-2'-deoxyadenosine (CdA) was found to be an effici
ent substrate for dGK (K(m), 85 muM). Subcellular fractionation of hum
an CEM lymphoblasts showed that extracts of mitochondria contain signi
ficant CdA phosphorylating activity (71.5 pmol/mg/min) that is not inh
ibited by excess of 2'-deoxycytidine (dCyd). This contrasts with the C
dA phosphorylating activity found in cytosolic extracts, which is carr
ied out by dCyd kinase and strongly inhibited by excess of dCyd. The e
fficient CdA phosphorylation by mitochondrial dGK is a novel finding t
hat may have far reaching implications for the clinical use of this po
tent cytostatic drug.