Section 6
Chapter 5,707

Inhibition of purified human and herpes simplex virus-induced DNA polymerases by 9- (2-hydroxyethoxymethyl) guanine triphosphate. Effects on primer-template function

Derse, D.; Cheng, Y.C.; Furman, P.A.; St Clair, M.H.; Elion, G.B.

Journal of Biological Chemistry 256(22): 11447-11451


ISSN/ISBN: 0021-9258
PMID: 6271750
Accession: 005706689

Download citation:  

The inhibition of highly purified herpes simplex virus (HSV)-induced and host cell DNA polymerases by the triphosphate form of 9-(2-hydroxyethoxymethyl)guanine (acyclovir; acycloguanosine) was examined. Acyclovir triphosphate (acyclo-GTP) competitively inhibited the incorporation of dGMP into DNA catalyzed by HSV DNA polymerase; apparent Km and Ki values of dGTP and acyclo-GTP were 0.15 .mu.M and 0.003 .mu.M, respectively. HelA [human cervical carcinoma cell] DNA polymerase .alpha. was also competitively inhibited; Km and Ki values of dGTP and acyclo-GTP were 1.2 and 0.18 .mu.M, respectively. HeLa DNA polymerase .beta. was insensitive to th analog. The limited DNA synthesis observed when dGTP was omitted from HSV or .alpha. DNA polymerase reactions was inhibited by acyclo-GTP in a concentration-dependent manner. Prior incubation of activated DNA, acyclo-GTP and DNA polymerase (.alpha. or HSV) resulted in a marked decrease in the utilization of the primer-template in subsequent DNA polymerase reactions. This decreased ability of preincubated primer-templates to support DNA synthesis w4s dependent on acyclo-GTP, enzyme concentration and the time or prior incubation. Acyclo-GMP-terminated DNA inhibited HSV DNA polymerase-catalyzed DNA synthesis. Kinetic experiments with variable concentrations of activated DNA and fixed concentrations of acyclo-GMP-terminated DNA revealed a noncompetitive inhibition of HSV-1 DNA polymerase. The apparent Km of 3'-hydroxyl termini was 1.1 .times. 10-7 M, the Kii and Kis of acyclo-GMP termini in activated DNA were 8.8 .times. 10-8 M and 2.1 .times. 10-9 M, respectively. 14C-labeled acylco-GMP residues incorporated ito activated DNA by HSV-1 DNA polymerase could not be excised by the polymerase-associated 3',5'-exonuclease activity.

PDF emailed within 1 workday: $29.90