Mutagenesis of conserved region I in the DNA polymerase from human adenovirus serotype 2.
The functional importance of the conserved region I (YGDTDSLF) found in several prokaryotic, eukaryotic, and viral DNA polymerases has been probed by site-directed mutagenesis of the adenovirus DNA polymerase (Ad Pol). Three different adenovirus-specific assays have been used to measure the in vitro activity of region I mutants of Ad Pol expressed in transiently transfected CMT-4 cells. In general, both conservative and nonconservative changes generally showed a greater than 5- to 10-fold reduction in activity in three different assays for activity. However, several replacements at the glycine residue showed activities closer to wild-type levels. For example, replacements of this glycine with cysteine (found in bacteriophage phi 29, another protein primed replication system), with serine, or with methionine had little effect on the activity observed in adenovirus-specific assays, such as initiation and elongation. These studies confirm the importance of this region of Ad Pol in specific initiation and elongation reactions on Ad DNA templates.
Joung
I
I
Department of Biochemistry, University of Alabama, Birmingham 35294.
Horwitz
M S
MS
Engler
J A
JA
D000260:Adenoviruses, Human / Q000201:enzymology / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D005075:Biological Evolution; D002460:Cell Line; D003593:Cytoplasm / Q000201:enzymology; D004261:DNA Replication; D004259:DNA-Directed DNA Polymerase / Q000235:genetics* / Q000378:metabolism; D005998:Glycine; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D012689:Sequence Homology, Nucleic Acid; D014162:Transfection
D004259:DNA-Directed DNA Polymerase; D005998:Glycine