Photochemical cross-linking has been used to identify residues in the Escherichia coli RecA protein that are proximal to and may directly mediate binding of DNA. Ultraviolet irradiation promotes specific and efficient cross-linking of the RecA protein to poly(deoxythymidylic) acid. Cross- linked peptides remaining covalently attached to the polynucleotide following proteolytic digestion with trypsin correspond to amino acids 61-72, 178-183, and 233-243 of the RecA protein primary sequence. Their location and surface accessibility in the crystal structure, along with the behavior of various recA mutants, support the assignment of the cross-linked regions to the DNA binding site(s) of the RecA protein. Functional overlap of amino acids 61-72 with an element of the ATP binding site suggests a structural mechanism by which nucleotide cofactors allosterically affect the RecA nucleoprotein filament.
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