Linker insertion mutagenesis of the human immunodeficiency virus reverse transcriptase expressed in bacteria

Definition of the minimal polymerase domain

Vinayaka R. Prasad, S. P. Goff

Research output: Contribution to journalArticle

82 Citations (Scopus)

Abstract

A plasmid construct expressing the p66 version of the human immunodeficiency virus reverse transcriptase as a bacterial fusion protein was subjected to in vitro mutagenesis, and the resulting variant proteins were assayed to define the locations of the two major enzymatic activities. The DNA polymerase activity was localized to the N-terminal portion of the protein; mutations altering or eliminating the C-terminal portion had little or no effect on that activity. The results suggest that, in contrast with previous reports, the p51 subunit found in virions should exhibit DNA polymerase activity. Mutations in many parts of the protein elimination RNAase H activity, suggesting that several areas are needed for proper folding and generation of that activity.

Original languageEnglish (US)
Pages (from-to)3104-3108
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume86
Issue number9
StatePublished - 1989
Externally publishedYes

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HIV Reverse Transcriptase
Insertional Mutagenesis
DNA-Directed DNA Polymerase
Bacteria
Ribonuclease H
Mutation
Proteins
Bacterial Proteins
Mutagenesis
Virion
Plasmids

ASJC Scopus subject areas

  • General
  • Genetics

Cite this

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AB - A plasmid construct expressing the p66 version of the human immunodeficiency virus reverse transcriptase as a bacterial fusion protein was subjected to in vitro mutagenesis, and the resulting variant proteins were assayed to define the locations of the two major enzymatic activities. The DNA polymerase activity was localized to the N-terminal portion of the protein; mutations altering or eliminating the C-terminal portion had little or no effect on that activity. The results suggest that, in contrast with previous reports, the p51 subunit found in virions should exhibit DNA polymerase activity. Mutations in many parts of the protein elimination RNAase H activity, suggesting that several areas are needed for proper folding and generation of that activity.

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