Turnover of endogenous SsrA-tagged proteins mediated by ATP-dependent proteases in Escherichia coli.
Formation and degradation of SsrA-tagged proteins enable ribosome recycling and elimination of defective products of incomplete translation. We produced an antibody against the SsrA peptide and used it to measure the amounts of SsrA-tagged proteins in Escherichia coli cells without interfering with tagging or altering the context of the tag added at the ends of nascent polypeptides. SsrA-tagged proteins were present in very small amounts unless a component of the ClpXP protease was missing. From the levels of tagged proteins in cells in which degradation is essentially blocked, we calculate that > or =1 in 200 translation products receives an SsrA tag. ClpXP is responsible for > or =90% of the degradation of SsrA-tagged proteins. The degradation rate in wild type cells is > or =1.4 min(-1) and decreases to approximately 0.10 min(-1) in a clpX mutant. The rate of degradation by ClpXP is decreased approximately 3-fold in mutants lacking the adaptor SspB, whereas degradation by ClpAP is increased 3-5-fold. However, ClpAP degrades SsrA-tagged proteins slowly even in the absence of SspB, possibly because of interference from ClpA-specific substrates. Lon protease degrades SsrA-tagged proteins at a rate of approximately 0.05 min(-1) in the presence or absence of SspB. We conclude that ClpXP, together with SspB, is uniquely adapted for degradation of SsrA-tagged proteins and is responsible for the major part of their degradation in vivo.
Laboratory of Cell Biology, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Mark
M
Lies
Michael R
MR
Maurizi
D000074183:ATPases Associated with Diverse Cellular Activities; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000737:chemistry*; D018407:Bacterial Physiological Phenomena; D002352:Carrier Proteins / Q000378:metabolism; D049071:Endopeptidase Clp / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D007700:Kinetics; D008954:Models, Biological; D018832:Molecular Chaperones / Q000378:metabolism; D009154:Mutation; D012329:RNA, Bacterial / Q000378:metabolism*
D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; D012329:RNA, Bacterial; C477700:sspB protein, E coli; C096222:tmRNA; D000255:Adenosine Triphosphate; C483568:ClpA protease, E coli; C483572:ClpP protease, E coli; D049071:Endopeptidase Clp; D000251:Adenosine Triphosphatases; C088072:ClpX protein, E coli; D000074183:ATPases Associated with Diverse Cellular Activities