Direct visualization of dynamics and co-operative conformational changes within RecA filaments that appear to be associated with the hydrolysis of adenosine 5'-O-(3-thiotriphosphate).

Abstract:

:Highly co-operative structural transitions and conformational changes can be directly observed in bundles of filaments formed by the RecA protein of Escherichia coli. These filaments have been formed with RecA protein, DNA and the ATP analog adenosine 5'-O-(3-thiotriphosphate) (ATP-gamma-S). We show that while ATP-gamma-S has frequently been called non-hydrolyzable in the RecA literature, it is hydrolyzed by RecA with a kcat of about 0.01 to 0.005 min-1. This rate of ATP-gamma-S hydrolysis is significant to structural studies conducted on a time scale of hours. It has been shown that RecA subunits may be seen in different conformations within one particular form of RecA bundle. We now show that additional structural transitions take place within these bundles when they are allowed to incubate at 37 degrees C for several hours. This is the same time scale on which ATP-gamma-S is being hydrolyzed, and the suggestion that the observed structural transitions arise from the hydrolysis of ATP-gamma-S is supported by the fact that when the hydrolysis of ATP-gamma-S is inhibited (at 4 degrees C), the transitions are not observed. The transitions that occur are highly co-operative, with filaments as a whole changing their state over lengths of several thousand Angstroms. This shows that RecA filaments have an internal co-operativity, and we suggest that this is important to their function in vivo. The motions of subunits that we visualize appear to be mainly rotational, and this can be used to infer information about the motions of RecA subunits associated with the RecA ATPase that occurs during the DNA strand exchange reaction.

journal_name

J Mol Biol

authors

Yu X,Egelman EH

doi

10.1016/0022-2836(92)91036-o

keywords:

subject

Has Abstract

pub_date

1992-05-05 00:00:00

pages

193-216

issue

1

eissn

0022-2836

issn

1089-8638

pii

0022-2836(92)91036-O

journal_volume

225

pub_type

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