title: Importance of heptameric ring integrity for activity of Escherichia coli ClpP.
abstract: Radiation target analysis has been used to identify the minimal functional unit for expression of activity of ClpP, the proteolytic component of the ATP-dependent ClpAP protease. Radiation target sizes determined for small peptide hydrolysis, for ClpA activated and nucleotide-activated oligopeptide cleavage, and for ClpA-activated ATP-dependent protein degradation were 154, 118, and 160 kDa, respectively. Thus, the hydrolytic activity of ClpP, subunit M, 21,500, is dependent on the native oligomeric structure. The quaternary structure of ClpP determined by electron microscopy and hydrodynamic studies consists of two face-to-face seven-membered rings. The radiation target sizes are consistent with a requirement for conformational integrity of an entire ring for expression of hydrolytic activity. Radiation damage led to disruption of inter-ring contacts, giving rise to isolated rings of ClpP. Thus, contacts between rings of ClpP are less stable and more easily disrupted than contacts between subunits within the rings. Our data suggest that cooperative interactions between subunits within the ClpP rings are important for maintaining the active conformation of the proteolytic active site.
authors.affiliation: Laboratory of Structural Biology Research, National Institute of Arthritis and Musculoskeletal and Skin Diseases, Bethesda, Maryland 20892, USA.
authors.forename: M W
authors.initials: MW
authors.lastname: Thompson
authors.forename: J
authors.initials: J
authors.lastname: Miller
authors.forename: M R
authors.initials: MR
authors.lastname: Maurizi
authors.forename: E
authors.initials: E
authors.lastname: Kempner
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000378:metabolism* / Q000528:radiation effects; D000595:Amino Acid Sequence; D049071:Endopeptidase Clp; D004789:Enzyme Activation / Q000528:radiation effects; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D011487:Protein Conformation / Q000528:radiation effects; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000378:metabolism* / Q000528:radiation effects