Loading…

Characterisation of Bacillus stearothermophilus PcrA helicase: Evidence against an active rolling mechanism

PcrA from Bacillus stearothermophilus is a DNA helicase for which, despite the availability of a crystal structure, there is very little biochemical information. We show that the enzyme has a broad nucleotide specificity, even being able to hydrolyse ethenonucleotides, and is able to couple the hydr...

Full description

Saved in:
Bibliographic Details
Published in:Nucleic acids research 1998-06, Vol.26 (11), p.2686-2693
Main Authors: Bird, L. E., Brannigan, J. A., Subramanya, H. S., Wigley, D. B.
Format: Article
Language:English
Subjects:
Citations: Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:PcrA from Bacillus stearothermophilus is a DNA helicase for which, despite the availability of a crystal structure, there is very little biochemical information. We show that the enzyme has a broad nucleotide specificity, even being able to hydrolyse ethenonucleotides, and is able to couple the hydrolysis to unwinding of DNA substrates. In common with the Escherichia coli helicases Rep and UvrD, PcrA is a 3′–5′ helicase but at high protein concentrations it can also displace a substrate with a 5′ tail. However, in contrast to Rep and UvrD, we do not see any evidence for dimerisation of the protein even in the presence of DNA. The enzyme shows a specificity for the DNA substrate in gel mobility assays, with the preferred substrate being one with both single and double stranded regions of DNA. We propose that these data, together with existing structural evidence, support an inchworm rather than a rolling model for 3′–5′ helicase activity.
ISSN:0305-1048
1362-4962
1362-4962
DOI:10.1093/nar/26.11.2686