The Physarum polycephalum php gene encodes a unique cold-adapted serine-carboxyl peptidase, physarolisin II

Abstract 

The php gene from a true slime mold, Physarum polycephalum, is a late-replicating and transcriptionally active gene. The deduced amino acid sequence of the gene product is homologous to those of the serine-carboxyl peptidase family, including physarolisin I from the same organism, but lacks the propeptide region. In this study, the protein was expressed in Escherichia coli and shown to possess endopeptidase activity with unique substrate specificity. Thus, we named it physarolisin II. The enzyme was revealed to be a kind of cold-adapted enzyme since it was maximally active at 16–22°C. The active enzyme was markedly unstable due to rapid autolysis (t_1/2=∼5 min, at 18°C). At higher temperature, the enzyme was less active but more stable, despite the fact that no gross conformational change was observed by circular dichroism spectroscopy.

Keywords:  Autolysis, Cold-adapted enzyme, Physarolisin II, Serine-carboxyl peptidase, True slime mold, Physarum polycephalum

Abbreviations:  DAN, diazoacetyl-D,L-norleucine methyl ester, DFP, diisopropylfluorophosphate, EGTA, ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid, KPIEFF(NO₂)RL, Lys-Pro-Ile-Glu-Phe-Phe(4-NO₂)-Arg-Leu