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Cells of Escherichia coli selectively degrade proteins that have incorporated amino acid analogs. Within 1 hour after exposure of cells to canavanine, 50% of the analog-containing proteins were degraded to acid-soluble form. At the same time, no net loss of canavanine-containing protein occurred from the 100,000 X g supernatant. Instead, most of the proteins containing the analog, unlike normal ones, accumulated in particulate fractions sedimenting at 10,000 X g or 100,000 X g. They were then lost from these fractions concomitant with the degradation of the abnormal proteins. The loss of such proteins from particulate fractions accounted for all of the protein degraded to acid-soluble form. Similar observations were obtained after incorporation of other analogs or puromycin. The 10,000 X g pellets correspond to amorphous dense intracellular granules visible in electron micrographs of cells exposed to canavanine. Upon removal of the analog, these granules disappeared, simultaneously with the degradation of the analog-containing proteins. These pellets do not resemble a degradative organelle, like the lysosome; they are not osmotically sensitive, do not exclude inulin, are not enclosed by a membrane, and do not show autolytic activity. The proteins in the granules could be solubilized by sodium dodecyl sulfate but not by Triton, NaC1, dithiothreitol, RNase, DNase, or phospholipase. The proteins extracted from the pellet with sodium dodecyl sulfate tend to become particulate again upon removal of this detergent. Incorporation of canavanine caused a normally soluble polypeptide, the monomer of beta-galactosidase, to be inactive and found in the sedimentable fraction. These findings suggest that (a) the presence of amino acid analogs in proteins can make them less soluble, and (b) the inclusions are formed by the spontaneous precipitation of abnormal proteins rather than by an active granule-forming process.
Prouty et al. (Sat,) studied this question.