Abstract
We purified purple acid phosphatase (PAPase) from kidney bean (Phaseolus vulgaris L. Ohfuku) seeds, a Japanese cultivar of the kidney bean. The molecular mass of the purified native enzyme was estimated approximately 110kDa by gel filtration. Following SDS-PAGE in the presence of 2-mercaptoethanol, glycosylated polypeptides of major 61kDa and minor 59kDa were observed. The N-terminal amino acid sequences for both bands were determined to be GKSSNFVRKTNKNRDMPLDS, suggesting they were two different subunits with the same N-terminal but probably different C-terminal or degree of glycosylation. The partial nucleotide sequences covering the N-terminal region were also determined using polymerase chain reaction (PCR). Comparison with the deduced amino acid sequence from the nucleotide sequence and that of the purified enzyme revealed that the N-terminal amino acid residue corresponded to Gly 23 from the initiation colon (Met). For the deduced amino acid sequence of the PAPase gene, a SignalP (ver. 2.0) analysis program predicted that a 22 amino acid sequence at the N-terminus of a precursor protein was signal peptide. This prediction was in agreement with the fact that PAPase with a signal peptide at the N-terminus was cleaved between Gly 22 and Gly 23 by putative signal peptidase.
