Inhibition of Eschericia coli RecA by a rationally designed peptide helix

BIOL 230

Daniel J Cline, clined@email.unc.edu, School of Pharmacy--Division of Medicinal Chemistry & Natural Products, University of North Carolina at Chapel Hill, 304c Beard Hall, Chapel Hill, NC 27599 and Scott F Singleton, sfs@email.unc.edu, Division of Medicinal Chemistry and Natural Products, School of Pharmacy, University of North Carolina, CB # 7360, Beard Hall 304C, Chapel Hill, NC 27599-7360.
Bacterial RecA functions as an ATP-dependent filamentous homopolymer assembled around ssDNA and promotes SOS response to DNA damage and homologous recombination, which may confer antibiotic resistance. A 29mer peptide was designed from the RecA N-terminal domain involved in inter-monomer contact, incorporating stable secondary structure elements and a reactive cysteine residue. This peptide inhibits the RecA's ATPase activity with an IC50 of 33 µM under reducing conditions. Under non-reducing conditions, 10 µM peptide is sufficient to inhibit completely. An additional derivative with a reactivity-enhanced Cys completely inhibited RecA at 5 µM, while another with an acylated Cys behaved similarly to the original peptide under reducing conditions. A gel shift of inhibited RecA was observed and was reversed by reductant. Disulfide-bonded chymotrypsin fragments were identified by in-gel digestion and MALDI-MS. These peptides provide the first step towards the design of first generation small-molecule inhibitors of RecA activities.
 

Enzymes
4:30 PM-6:30 PM, Wednesday, 13 September 2006 Moscone Center -- Hall D, Poster

Division of Biological Chemistry

The 232nd ACS National Meeting, San Francisco, CA, September 10-14, 2006