Limits...
Unwinding forward and sliding back: an intermittent unwinding mode of the BLM helicase.

Wang S, Qin W, Li JH, Lu Y, Lu KY, Nong DG, Dou SX, Xu CH, Xi XG, Li M - Nucleic Acids Res. (2015)

Bottom Line: Mutational studies revealed that the RQC domain plays an important role in stabilizing the helicase/DNA interaction during both DNA unwinding and backward sliding of BLM.Especially, Lys1125 in the RQC domain, a highly conserved amino acid among RecQ helicases, may be involved in the backward sliding activity.These results may shed new light on the mechanisms for BLM in DNA repair and homologous recombination.

View Article: PubMed Central - PubMed

Affiliation: Beijing National Laboratory for Condensed Matter Physics and CAS Key Laboratory of Soft Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China.

Show MeSH

Related in: MedlinePlus

Characterizations of the DNA binding and unwinding activities of the wild-type and mutant BLM. The polarization anisotropy assay for DNA binding and stopped-flow assay for DNA unwinding were performed as described in the Supplementary data. (A and B) Binding curves for ssDNA (18 nt) (A) and dsDNA (18 bp) (B). The dissociation constants were obtained from fits with the Hill equation and given in Table 1. (C) Kinetic DNA unwinding curves.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4402530&req=5

Figure 4: Characterizations of the DNA binding and unwinding activities of the wild-type and mutant BLM. The polarization anisotropy assay for DNA binding and stopped-flow assay for DNA unwinding were performed as described in the Supplementary data. (A and B) Binding curves for ssDNA (18 nt) (A) and dsDNA (18 bp) (B). The dissociation constants were obtained from fits with the Hill equation and given in Table 1. (C) Kinetic DNA unwinding curves.

Mentions: DNA binding measurements showed that these mutants have similar binding affinities for ssDNA as the wild-type enzyme (Figure 4A and Table 1), whereas their binding affinities for dsDNA are more significantly reduced (FigureĀ 4B and Table 1). From the dissociation constants, the binding affinities for dsDNA are in the following order: wilde-type > S1121A > K1125A > T1110G. DNA unwinding efficiencies from further stopped-flow kinetic measurements essentially agree with the binding results, indicating that the RQC domain plays an important role in the DNA unwinding activity of BLM.


Unwinding forward and sliding back: an intermittent unwinding mode of the BLM helicase.

Wang S, Qin W, Li JH, Lu Y, Lu KY, Nong DG, Dou SX, Xu CH, Xi XG, Li M - Nucleic Acids Res. (2015)

Characterizations of the DNA binding and unwinding activities of the wild-type and mutant BLM. The polarization anisotropy assay for DNA binding and stopped-flow assay for DNA unwinding were performed as described in the Supplementary data. (A and B) Binding curves for ssDNA (18 nt) (A) and dsDNA (18 bp) (B). The dissociation constants were obtained from fits with the Hill equation and given in Table 1. (C) Kinetic DNA unwinding curves.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4402530&req=5

Figure 4: Characterizations of the DNA binding and unwinding activities of the wild-type and mutant BLM. The polarization anisotropy assay for DNA binding and stopped-flow assay for DNA unwinding were performed as described in the Supplementary data. (A and B) Binding curves for ssDNA (18 nt) (A) and dsDNA (18 bp) (B). The dissociation constants were obtained from fits with the Hill equation and given in Table 1. (C) Kinetic DNA unwinding curves.
Mentions: DNA binding measurements showed that these mutants have similar binding affinities for ssDNA as the wild-type enzyme (Figure 4A and Table 1), whereas their binding affinities for dsDNA are more significantly reduced (FigureĀ 4B and Table 1). From the dissociation constants, the binding affinities for dsDNA are in the following order: wilde-type > S1121A > K1125A > T1110G. DNA unwinding efficiencies from further stopped-flow kinetic measurements essentially agree with the binding results, indicating that the RQC domain plays an important role in the DNA unwinding activity of BLM.

Bottom Line: Mutational studies revealed that the RQC domain plays an important role in stabilizing the helicase/DNA interaction during both DNA unwinding and backward sliding of BLM.Especially, Lys1125 in the RQC domain, a highly conserved amino acid among RecQ helicases, may be involved in the backward sliding activity.These results may shed new light on the mechanisms for BLM in DNA repair and homologous recombination.

View Article: PubMed Central - PubMed

Affiliation: Beijing National Laboratory for Condensed Matter Physics and CAS Key Laboratory of Soft Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China.

Show MeSH
Related in: MedlinePlus