Recognition of 2'-deoxy-l-ribonucleoside 5'-triphosphates by human telomerase.
Telomerase is classified as one of the reverse transcriptases (RTs). To clarify whether l-enantiomers of natural 2'-deoxyribonucleoside 5'-triphosphates (dNTPs) are recognized by human telomerase, a quantitative telomerase assay based on the "stretch PCR" method was developed and used for kinetic analysis of the inhibitory effects of these compounds on the enzyme. Among the four l-enantiomers of dNTPs, l-dTTP and l-dGTP inhibited telomerase activity and the others showed slight or no inhibitory effect. Lineweaver-Burk plot analysis showed that the inhibition modes of l-dTTP and l-dGTP were partially competitive (mixed type) and competitive with the corresponding substrate dNTP, respectively. However, the K(i) values of l-dTTP and l-dGTP (21 and 15 microM) were several times larger than the K(m) values (3-6 microM). These results suggest that the active site of telomerase is not able to discriminate strictly the chirality of dNTPs, although it is more discriminatory than HIV-1 RT.
Yamaguchi
T
T
Department of Biological Sciences, Teikyo University of Science and Technology, Uenohara, Yamanashi, 409-0193, Japan. toyofumi@ntu.ac.jp
Yamada
R
R
Tomikawa
A
A
Shudo
K
K
Saito
M
M
Ishikawa
F
F
Saneyoshi
M
M
D003838:Deoxyadenine Nucleotides / Q000737:chemistry / Q000378:metabolism; D003845:Deoxycytosine Nucleotides / Q000737:chemistry / Q000378:metabolism; D003848:Deoxyguanine Nucleotides / Q000737:chemistry / Q000378:metabolism; D003854:Deoxyribonucleotides / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D013237:Stereoisomerism; D013379:Substrate Specificity; D019098:Telomerase / Q000378:metabolism*; D013942:Thymine Nucleotides / Q000737:chemistry / Q000378:metabolism
D003838:Deoxyadenine Nucleotides; D003845:Deoxycytosine Nucleotides; D003848:Deoxyguanine Nucleotides; D003854:Deoxyribonucleotides; D013942:Thymine Nucleotides; C024107:2'-deoxycytidine 5'-triphosphate; C029603:deoxyguanosine triphosphate; D019098:Telomerase; C026600:2'-deoxyadenosine triphosphate; C024157:thymidine 5'-triphosphate