Purification and characterization of the carboxypeptidase isoinhibitors from potatoes.

Three zones of carboxypeptidase inhibitory activity were observed when heat-stable extracts of potato tubers (cv. Russet Burbank) were chromatographed on carboxymethyl cellulose. The isoinhibitors found in these zones were denoted I, II, and III based upon their order of elution from this column. The predominant form (II) had previously been suggested to be a mixture of two polypeptides (IIa and IIb) differing in that IIa possessed an additional residue of glutamine (Hass et al. 1975 Biochemistry 14: 1334). These closely related isoinhibitors (IIa and IIb) were separated by equilibrium ion exchange chromatography and characterized. Isoinhibitor I was shown to be identical to II except for two replacements, Ser-30 --> Ala and Arg-32 --> Gly. These replacements had no significant effect on apparent K(i) values toward either carboxypeptidase A or B. Isoinhibitor III, which was identical to II except that it lacked the amino terminal pyrrolidone carboxylic acid and following glutamine residue, was also functionally indistinguishable from II in inhibition studies. It was concluded that at least two and possibly as many as five genes code for the various isoinhibitor species which are present in potato tubers.