Phosphorylated peptides can limit Saccobolus platensis aminopeptidase action

Abstract

The effect of substrate phosphorylation on the susceptibility to proteolytic cleavage by purified aminopeptidase from Saccobolus platensis was investigated using the model heptapeptide L-R-R-A-S-L-G. Phosphorylation of serine greatly altered the action of peptidase producing a fragment, A-S(P)-L-G, insensitive to further attack by the peptidase. The action of peptidase was tested on peptides generated by subtilisin digestion of fungal cytosolic proteins labeled in vivo with [³H]leucine and phosphorylated in vitro with the catalytic subunit of cyclic AMP-dependent protein kinase. Phosphopeptides were enriched by gel filtration through P-2 columns. After exhaustive exopeptidase degradation the peak of [³²p]phosphopeptides remained mostly unchanged. Removal of phosphate with alkaline phosphatase prior to treatment with peptidase produced a 12% liberation of [³H]leucine. The results support the idea that phosphorylation influences final protein processing.