Functional roles of carboxylate residues comprising the DNA polymerase active site triad of Ty3 reverse transcriptase.
Functional roles of carboxylate residues comprising the DNA polymerase active site triad of Ty3 reverse transcriptase.
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DOI：
10.1093/nar/gki150
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发表时间：
2005
影响因子：
14.9
通讯作者：
Le Grice SF
中科院分区：
文献类型：
--
作者：
Bibillo A;Lener D;Klarmann GJ;Le Grice SF
Aspartic acid residues comprising the -D-(aa)n-Y-L-D-D- DNA polymerase active site triad of reverse transcriptase from the Saccharomyces cerevisiae long terminal repeat-retrotransposon Ty3 (Asp151, Asp213 and Asp214) were evaluated via site-directed mutagenesis. An Asp151→Glu substitution showed a dramatic decrease in catalytic efficiency and a severe translocation defect following initiation of DNA synthesis. In contrast, enzymes harboring the equivalent alteration at Asp213 and Asp214 retained DNA polymerase activity. Asp151→Asn and Asp213→Asn substitutions eliminated both polymerase activities. However, while Asp214 of the triad could be replaced by either Asn or Glu, introducing Gln seriously affected processivity. Mutants of the carboxylate triad at positions 151 and 213 also failed to catalyze pyrophosphorolysis. Finally, alterations to the DNA polymerase active site affected RNase H activity, suggesting a close spatial relationship between these N- and C-terminal catalytic centers. Taken together, our data reveal a critical role for Asp151 and Asp213 in catalysis. In contrast, the second carboxylate of the Y-L-D-D motif (Asp214) is not essential for catalysis, and possibly fulfills a structural role. Although Asp214 was most insensitive to substitution with respect to activity of the recombinant enzyme, all alterations at this position were lethal for Ty3 transposition.
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影响因子：
2.9
作者：
Ghosh, M;Jacques, PS;leGrice, SFJ
通讯作者：
leGrice, SFJ
DOI：
10.1073/pnas.90.13.6320
发表时间：
1993-07-01
影响因子：
11.1
作者：
JACOBOMOLINA, A;DING, JP;ARNOLD, E
通讯作者：
ARNOLD, E
影响因子：
5.4
作者：
HANSEN, LJ;SANDMEYER, SB
通讯作者：
SANDMEYER, SB
影响因子：
5.3
作者：
HANSEN, LJ;CHALKER, DL;SANDMEYER, SB
通讯作者：
SANDMEYER, SB
影响因子：
3.7
作者：
Boyer, PL;Gao, HQ;Hughes, SH
通讯作者：
Hughes, SH