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. Author manuscript; available in PMC: 2010 Mar 20.

Published in final edited form as: Cell. 2009 Mar 20;136(6):1032–1043. doi: 10.1016/j.cell.2009.02.019

(A) Autoradiograph of an agarose gel showing RAD51 (3 μM) incubated with either GST-BRC4 or GST-Δ1524-30 prior to incubation with ³²P-labeled ϕX174 linear dsDNA (5 μM, nt) and further incubation for 1 h in the presence of ATP, Mg²⁺ and Ca²⁺. (B) Quantification of data as in (A): in the presence of ATP, Mg²⁺ + Ca²⁺ (●); ADP, Mg²⁺ + Ca²⁺ (); ATP and Ca²⁺ (); or AMP-PNP, Mg²⁺ + Ca²⁺ (). Filled circles and solid lines correspond to GST-BRC4; open circles and dashed lines correspond to GST-Δ1524-30. Data obtained with the control mutant, GST-T1526A, in the presence of ATP, Mg²⁺ + Ca²⁺ :(). (C) The effect of BRC4 or BRCA2_BRC1-8 on RAD51-dsDNA complex formation using dA₄₀·dT₄₀ dsDNA. RAD51 (0.15 μM) was incubated with GST-BRC4 or BRCA2_BRC1-8, at the concentrations indicated prior to incubation with ³²P-labeled dA₄₀·dT₄₀ dsDNA (2.4 μM, nt) and further incubation for 1 h in the presence of ATP, Mg²⁺ and Ca²⁺. Protein-DNA complexes were resolved in 6% PAGE and analyzed by autoradiography. (D) Data obtained as in (C), quantified and plotted: GST-BRC4, (●); BRCA2_BRC1-8 ().

Inline graphic — (A) Autoradiograph of an agarose gel showing RAD51 (3 μM) incubated with either GST-BRC4 or GST-Δ1524-30 prior to incubation with ³²P-labeled ϕX174 linear dsDNA (5 μM, nt) and further incubation for 1 h in the presence of ATP, Mg²⁺ and Ca²⁺. (B) Quantification of data as in (A): in the presence of ATP, Mg²⁺ + Ca²⁺ (●); ADP, Mg²⁺ + Ca²⁺ (); ATP and Ca²⁺ (); or AMP-PNP, Mg²⁺ + Ca²⁺ (). Filled circles and solid lines correspond to GST-BRC4; open circles and dashed lines correspond to GST-Δ1524-30. Data obtained with the control mutant, GST-T1526A, in the presence of ATP, Mg²⁺ + Ca²⁺ :(). (C) The effect of BRC4 or BRCA2_BRC1-8 on RAD51-dsDNA complex formation using dA₄₀·dT₄₀ dsDNA. RAD51 (0.15 μM) was incubated with GST-BRC4 or BRCA2_BRC1-8, at the concentrations indicated prior to incubation with ³²P-labeled dA₄₀·dT₄₀ dsDNA (2.4 μM, nt) and further incubation for 1 h in the presence of ATP, Mg²⁺ and Ca²⁺. Protein-DNA complexes were resolved in 6% PAGE and analyzed by autoradiography. (D) Data obtained as in (C), quantified and plotted: GST-BRC4, (●); BRCA2_BRC1-8 ().