Open AccessNucleotide binding by a 24-residue peptide from the RecA protein of Escherichia coli.
Author(s) -
Kendall L. Knight,
Kevin McEntee
Publication year - 1986
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.83.24.9289
Subject(s) - peptide , escherichia coli , residue (chemistry) , chemistry , biochemistry , photoaffinity labeling , adenosine triphosphate , affinity label , nucleotide , tyrosine , binding site , gene
We have recently demonstrated that two ATP analog affinity labels, 8-azidoadenosine 5'-triphosphate (N3ATP) and 5'-p-fluorosulfonylbenzoyladenosine (5'FSBA), covalently modify RecA protein of Escherichia coli at a specific tyrosine residue (Tyr-264) located within a 24-residue tryptic peptide (T-31) spanning residues 257-280. Here we show that N3ATP efficiently modifies purified peptide T-31 and show that the interaction is specific by the following criteria: photolabeling of peptide T-31 is saturable with respect to the N3ATP concentration; photolabeling is competitive with ATP and adenosine but not with adenine, UTP, or TTP; and other peptides derived from RecA protein were poor substrates for photolabeling except for one fragment that showed a nonspecific interaction with the photoaffinity analog. Analysis of N3ATP-modified T-31 shows that the photolabel attaches to more than one site within the peptide. These data argue that peptide T-31 contains some sites of contact for adenine and ribose moieties of ATP when it is bound to RecA protein.