Ornithine decarboxylase (ODC) can be induced up to 100-fold over basal levels four hours after addition of glutamine to the medium of HeLa cells growing in suspension culture. As demonstrated in several other cell types, ODC is inactivated very rapidly in HeLa cells, and the rate of inactivation is seen to vary with a half life of 9-15 minutes in uninduced cells and rises to ca. 60 minutes at the peak of induction. Quantitatively, the change in rate of inactivation cannot completely account for the observed rise in activity, thus synthesis or activation of ODC must also be involved in the induction process. The inactivation process requires metabolic energy and it can be sustained by glycolytic derived energy. Other factors which are known to inhibit protein breakdown in mammalian cells, such as sodium fluoride, insulin, or tosyl phenylalanyl chloromethyl ketone, had no effect on the rate of inactivation of ODC. Attempts to demonstrate ODC inactivation in a cell free system at neutral pH were unsuccessful.

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A general method for obtaining high-level production of low molecular weight proteins in Escherichia coli is described. This method is based on the use of a novel Met-X,,-protein construction which is formed by insertion of a single amino acid residue (preferably Arginine or Lysine) between the N-terminal methionine and the protein of interest. The utility of this method is illustrated by examples for achieving high-level production of human insulin-like growth factor-1, human proinsulin, and their analogs. Furthermore, highly produced insulin-like growth factor-] derivatives and human proinsulin analogs are converted to their natural sequences by removal of dipeptides with cathepsin C.

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