Related functional domains in virus DNA polymerases.

Abstract

Analysis of the lesions in several drug-resistant DNA polymerase mutants of herpes simplex virus along with comparative analysis of the published polymerase sequences of other human herpesviruses has shown that most lesions (five out of six) are substitutions at amino acid residues conserved in all four polymerases. Furthermore, the majority of lesions are in regions of the polypeptide where there are marked clusterings of conserved residues. On the basis of these data we have identified several domains within the polypeptide which we believe may have important functional roles in the action of the enzyme. The apparent restriction in the potential sites of lesions conferring drug resistance may explain the difficulty in selecting such mutants using acyclovir (ACV) in culture and their failure to emerge so far during ACV therapy. Extension of the comparative analysis to the polymerases of adenovirus type 2, vaccinia virus and phage q29 suggests that these enzymes also possess domains homologous to those most conserved in the herpes polymerases (regions I-HI) and that these domains have a similar linear spatial distribution on the polypeptides. The results are discussed in relation to the known function of the DNA polymerases.

Characterization of a DNA Polymerase Mutant of Herpes Simplex Virus from a Severely Immunocompromised Patient Receiving Acyclovir

“…This change would lead to a predicted amino acid substitution (Gly ~ Set) at residue 841. The change observed in 8-H9 was identical to that found previously in a laboratory-generated DNA polymerase mutant (TP2.7) derived from the wild-type SC16 (Larder et al, 1987). In this case it was confirmed by marker rescue analysis that the mutation in TP2.7 conferred resistance to ACV.…”

“…The cloning strategy described by Larder et al (1987) was used to obtain fragments of the DNA polymerase genes for sequencing. Procedures used for ssDNA sequencing have also been published previously (Larder et al, 1987).…”

“…The cloning strategy described by Larder et al (1987) was used to obtain fragments of the DNA polymerase genes for sequencing. Procedures used for ssDNA sequencing have also been published previously (Larder et al, 1987). Briefly, ssDNA was isolated by polyethylene glycol precipitation from M13mp8 clones and inserts were sequenced by the dideoxynucleotide chain termination method of Sanger et al (1977).…”

“…The polymerase gene of the laboratory wild-type HSV-1 strain (SC16) has been sequenced previously (Larder et al, 1987) and comparison of the polymerase gene of the ACV-sensitive isolate l-H2 with this laboratory strain revealed seven nucleotide changes leading to four amino acid substitutions. The extent of the sequence conservation observed between these strains is similar to that observed between other wild-type HSV-1 polymerase genes (Gibbs et al, 1985;Quinn & McGeoch, 1985;Knopf, 1987 A number of HSV-1 isolates obtained from a bone marrow transplant patient receiving ACV prophylaxis and subsequent therapy for indolent oral HSV, were found to be resistant to ACV.…”

Recent advances in the molecular biology of entomopoxviruses

“…It has been previously established that there are conserved regions shared by pol genes from several viruses of vertebrates and prokaryotes. Four of these regions seem to be more highly conserved than others Gibbs et al, 1985;Larder et al, 1987;Mustafa & Yuen, 1991;Tomalski et al, 1988). In these four regions there is 32-46% amino acid identity between the DNA polymerase of CbEPV and that of vaccinia virus.…”