The ClpP peptidase is the major determinant of bulk protein turnover in Bacillus subtilis.
Measurements of overall protein degradation rates in wild-type and clpP mutant Bacillus subtilis cells revealed that stress- or starvation-induced bulk protein turnover depends virtually exclusively on the ClpP peptidase. ClpP is also essential for intracellular protein quality control, and in its absence newly synthesized proteins were highly prone to aggregation even at 37 degrees C. Proteomic comparisons between the wild type and a DeltaclpP mutant showed that the absence of ClpP leads to severe perturbations of "normal" physiology, complicating the detection of ClpP substrates. A pulse-chase two-dimensional gel approach was therefore used to compare wild-type and clpP mutant cultures that had been radiolabeled in mid-exponential phase, by quantifying changes in relative spot intensities with time. The results showed that overall proteolysis is biased toward proteins with vegetative functions which are no longer required (or are required at lower levels) in the nongrowing state. The identified substrate candidates for ClpP-dependent degradation include metabolic enzymes and aminoacyl-tRNA synthetases. Some substrate candidates catalyze the first committed step of certain biosynthetic pathways. Our data suggest that ClpP-dependent proteolysis spans a broad physiological spectrum, with regulatory processing of key metabolic components and regulatory proteins on the one side and general bulk protein breakdown at the transition from growing to nongrowing phases on the other.
Ernst-Moritz-Arndt-Universität, Institut für Mikrobiologie, F-L-Jahn-Str. 15, 17487 Greifswald, Germany.
Holger
H
Kock
Ulf
U
Gerth
Michael
M
Hecker
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism; D001412:Bacillus subtilis / Q000201:enzymology / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D049071:Endopeptidase Clp; D004798:Enzymes / Q000378:metabolism; D017353:Gene Deletion; D005798:Genes, Bacterial; D009154:Mutation; D020543:Proteome / Q000032:analysis; D012697:Serine Endopeptidases / Q000378:metabolism*; D013194:Staining and Labeling; D013379:Substrate Specificity; D013462:Sulfur Radioisotopes / Q000378:metabolism
D001426:Bacterial Proteins; D004798:Enzymes; D020543:Proteome; D013462:Sulfur Radioisotopes; D012697:Serine Endopeptidases; D049071:Endopeptidase Clp; D000251:Adenosine Triphosphatases; D000604:Amino Acyl-tRNA Synthetases