Jiaa, H. and Korolev, S. and Niedziela-Majkaa, A. and Malufa, N.K. and Gaussa, G.H. and Myongf, S. and Hag, T. and Waksman, Gabriel and Lohman, T.M. (2011) Rotations of the 2B sub-domain of E. coli UvrD helicase/translocase coupled to nucleotide and DNA binding. Journal of Molecular Biology 411 (3), pp. 633-648. ISSN 0022-2836.Full text not available from this repository.
E. coli UvrD is a superfamily 1 (SF1) DNA helicase and single stranded (ss) DNA translocase that functions in DNA repair, plasmid replication and as an anti-recombinase by removing RecA protein from ssDNA. UvrD couples ATP binding and hydrolysis to unwind double-stranded DNA (dsDNA) and translocate along ssDNA with 3′ to 5′ directionality. Although a UvrD monomer is able to translocate along ssDNA rapidly and processively, DNA helicase activity in vitro requires a minimum of a UvrD dimer. Previous crystal structures of UvrD bound to a ss-duplex DNA junction show that its 2B sub-domain exists in a “closed” state and interacts with the duplex DNA. Here we report a crystal structure of an apo form of UvrD in which the 2B sub-domain is in an “open” state that differs by a ~160o rotation of the 2B sub-domain. To study the rotational conformational states of the 2B sub-domain in various ligation states, a series of double cysteine UvrD mutants were constructed and labeled with fluorophores such that rotation of the 2B sub-domain results in changes in fluorescence resonance energy transfer (FRET). These studies show that the open and closed forms can interconvert in solution with low salt favoring the closed conformation and high salt favoring the open conformation in the absence of DNA. Binding of UvrD to DNA as well as ATP binding and hydrolysis also affect the rotational conformational state of the 2B sub-domain suggesting that 2B sub-domain rotation is coupled to the function of this nucleic acid motor enzyme.
|Additional Information:||Corrigendum at http://dx.doi.org/10.1016/j.jmb.2012.02.030|
|Keyword(s) / Subject(s):||DNA repair, fluorescence, FRET, crystal structure, allostery|
|School or Research Centre:||Birkbeck Schools and Research Centres > School of Science > Biological Sciences|
|Date Deposited:||20 Jun 2011 08:32|
|Last Modified:||17 Apr 2013 12:20|
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