Crystal Structure of ClpP, the Proteolytic Component of Clp, the Chaperone-Linked Protease from E. coli. John M. Flanagan^*, James A. Hartling^*, Robert Sweet, Jimin Wang⁽, ^*Biology Dept. Brookhaven National Laboratory, Upton NY, 11973-5000, USA, ⁽Dept. Mol. Biochem. Biophys. Yale University, New Haven CT, 06511, USA

The crystal structure of E. coli ClpP, the serine protease component of the energy-dependent Clp protease, has been determined at 2.3 Å resolution. The 301-kilodalton proteolytic complex has 72-point group symmetry with its 14 subunits arranged in two stacked rings. A narrow channel controls access to a large central cavity containing the proteolytic active sites. The ClpP subunit has a previously unreported fold, although the architecture of its active sites shows similarities to those of other serine proteases. Two of the three residues in the catalytic triad, S97 and H122, lie in close proximity to a carboxylate, D171, which is the previously unidentified third member of the triad. The active sites lie along a hydrophobic channel at the edges of the oligomers large central cavity. In each ring, the active sites are spaced at ~28Å intervals along the channel, a distance that can be spanned by an extended hepta- or octa-peptide. The structure of ClpP in complex with the covalent inhibitor diisopropylflourophospate and also a peptide inhibitor suc-LLnL-al has also been determined. The significance of these structure to ATP-dependent protein degradation by Clp will also be discussed. The structure was solved using an ab initio phasing procedure exploiting the internal 14-fold symmetry of the oligomer. This approach may be useful in solving structures of other multimeric proteins.