Dynamic light scattering investigations of RecA self-assembly and interactions with single strand DNA

Benight, Albert S.; Wilson, David H.; Budzynski, Donna M.; Goldstein, Robert F.

doi:10.1016/0300-9084(91)90197-9

Cited by 16 publications

(15 citation statements)

References 30 publications

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“…These results are consistent with the notion of self-aggregation of hRAD51 in the absence of adenosine nucleotide. An aggregation effect that is competitive with ssDNA binding has been observed with RecA (18,19). In the case of hRAD51, the aggregate appears irreversible and is only likely to be relevant to in vitro assays.…”

Section: Discussionmentioning

confidence: 99%

Biochemical Characterization of the Human RAD51 Protein

Tombline

Heinen

Shim

et al. 2002

Journal of Biological Chemistry

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Adenosine nucleotides affect the ability of RecA⅐single-stranded DNA (ssDNA) nucleoprotein filaments to cooperatively assume and maintain an extended structure that facilitates DNA pairing during recombination. Here we have determined that ADP and ATP/ATP␥S affect the DNA binding and aggregation properties of the human RecA homolog human RAD51 protein (hRAD51). These studies have revealed significant differences between hRAD51 and RecA. In the presence of ATP␥S, RecA forms a stable complex with ssDNA, while the hRAD51 ssDNA complex is destabilized. Conversely, in the presence of ADP and ATP, the RecA ssDNA complex is unstable, while the hRAD51 ssDNA complex is stabilized. We identified two hRAD51⅐ssDNA binding forms by gel shift analysis, which were distinct from a well defined RecA⅐ssDNA binding form. The available evidence suggests that a low molecular weight hRAD51⅐ssDNA binding form (hRAD51⅐ssDNA low ) correlates with active ADP and ATP processing. A high molecular weight hRAD51⅐ ssDNA aggregate (hRAD51⅐ssDNA high ) appears to correlate with a form that fails to process ADP and ATP. Our data are consistent with the notion that hRAD51 is unable to appropriately coordinate ssDNA binding with adenosine nucleotide processing. These observations suggest that other factors may assist hRAD51 in order to mirror RecA recombinational function.

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Section: Discussionmentioning

confidence: 99%

Biochemical Characterization of the Human RAD51 Protein

Tombline

Heinen

Shim

et al. 2002

Journal of Biological Chemistry

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“…Although many AAA ϩ ATPases are homooligomeric, some are heterooligomeric and commonly contain ATPase active sites that are either functionally degenerate (i.e., Orc4 [26]) or inactive (processivity clamp loader [116]). In addition, an ATP-mediated subunit association, as suggested for Mcm2/5, is commonly observed for various viral helicases (i.e., SV40 TAg [259]), and gapped structures have been observed for other toroidal hexamers (i.e., SV40 TAg [21,175,176], RecA [15,240], and the Rho transcription terminator [234]). …”

Section: Model Of Mcm2-7 Functionmentioning

confidence: 91%

The Mcm Complex: Unwinding the Mechanism of a Replicative Helicase

Bochman

Schwacha

2009

Microbiol Mol Biol Rev

288

278

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SUMMARY The Mcm2-7 complex serves as the eukaryotic replicative helicase, the molecular motor that both unwinds duplex DNA and powers fork progression during DNA replication. Consistent with its central role in this process, much prior work has illustrated that Mcm2-7 loading and activation are landmark events in the regulation of DNA replication. Unlike any other hexameric helicase, Mcm2-7 is composed of six unique and essential subunits. Although the unusual oligomeric nature of this complex has long hampered biochemical investigations, recent advances with both the eukaryotic as well as the simpler archaeal Mcm complexes provide mechanistic insight into their function. In contrast to better-studied homohexameric helicases, evidence suggests that the six Mcm2-7 complex ATPase active sites are functionally distinct and are likely specialized to accommodate the regulatory constraints of the eukaryotic process.

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“…Analysis of the recombinational DNA repair and general genetic recombination activities of all mutants showed that only three of the wildtype Phe217,and Arg222) are important to these RecA functions, while the other 2 (Asn213 and Tyr218) are not (Skiba & Knight, 1994). The image in Figure 1 shows the proximity of residues 216, 217 and 222 (Brenner et al, 1988;Heuser & Grif®th, 1989; and by dynamic light scattering (Benight et al, 1991) (Benight et al, 1991) (Brenner et al, 1988;Heuser & Grif®th, 1989;Wilson & Benight, 1990; (Brenner et al, 1988;Heuser & Grif®th, 1989). Upon addition of nucleotide (0.5 mM ATPgS), bundles should be disrupted into the component ®laments, and longer ®laments will be reduced in size (Brenner et al, 1988;Heuser & Grif®th, 1989;Wilson & Benight, 1990 (Figure 2A).…”

Section: The Distribution Of These Different Forms Is a Function Ofmentioning

confidence: 93%