Article,

Protein Degradation in Escherichia coli. I. Measurement of rapidly and slowly decaying components

, and .
The Journal of Biological Chemistry, (June 1970)

Abstract

The rate of degradation of intracellular radioactive protein of bacterial cells supported by a membrane filter has been measured directly from the appearance of radioactivity in the carrier-containing perfusate. For this purpose a perfusion apparatus has been constructed from a microsyringe filter holder. The conditions inside the apparatus are physiological and permit exponential growth up to 1 x 108 cells. Exchange of extracellular carrier leucine-12C with the intracellular leucine-14C pool is so rapid that at least 87% of the amino acid arising from protein breakdown appears in the perfusate and is not recycled into protein even in growing cultures.Only a limited portion of the cellular protein is subject to rapid degradation. It decays with a half-life of approximately 1 hour and constitutes 2 to 7% of the total cellular protein of cells growing in glucose minimal medium under various nutritional conditions (including growth and starvation). The proportion of the total protein synthesis which is directed to the synthesis of this rapidly degrading protein component increases with decreasing growth rate. At very slow growth rates 10 to 40% of the radioactivity that is incorporated into protein is incorporated into the rapidly degrading protein. Treatment which removes nonproteinaceous material and Pronase digestion of cells before and after the decay of the rapid component substantiates the conclusion that the rapidly decaying component is derived from bacterial protein.There is another radioactive component in the perfusate which is released at the very slow rate of 0.2 to 0.6% per hour for as long as the experiments are carried out (48 hours). This does not represent degradation of intracellular protein to amino acids since this process is independent of leucine exchange, and 30 to 50% of the radioactivity in the perfusate is acid-insoluble.

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