Alteration by Site-directed Mutagenesis of the Conserved Lysine Residue in the Consensus ATP-binding Sequence of the RecB Protein of Escherichia Coli

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 1992 Nov 11

PMID 1454527

Citations 15

Authors

S Hsieh

D A Julin

Affiliations

Soon will be listed here.

Abstract

The RecB and RecD subunits of the RecBCD enzyme of Escherichia coli contain amino acid sequences similar to a consensus mononucleotide binding motif found in a large number of other enzymes. We have constructed by site-directed mutagenesis a lysine-to-glutamine mutation in this sequence in the RecB protein. The mutant enzyme (RecB-K29Q-CD) has essentially no nuclease or ATP hydrolysis activity on double-stranded DNA, showing the importance of RecB for unwinding double-stranded DNA. However, ATP hydrolysis stimulated by single-stranded DNA is reduced by only about 5-8-fold compared to the wild-type, nuclease activity on single-stranded DNA is reduced by less than 2-fold, and the nuclease activity of the RecB-K29Q-CD enzyme requires ATP. The effects of the RecB mutation suggest that the RecD protein hydrolyzes ATP and can stimulate the RecBCD enzyme nuclease activity on single-stranded DNA.

Citing Articles

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Amundsen S, Smith G J Mol Biol. 2024; 436(6):168482.

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E. coli RecB Nuclease Domain Regulates RecBCD Helicase Activity but not Single Stranded DNA Translocase Activity.

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Small-molecule sensitization of RecBCD helicase-nuclease to a Chi hotspot-activated state.

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