Cook M. & Adam Z. (1997). Purification and characterization of an arginyl peptidase from the chloroplast stroma of pea seedlings. Plant Physiol. Biochem. 35: 163-168.
Abstract
Stromal extract of pea (Pisum sativum var. Alaska)
chloroplasts was
fractionated by column chromatography, using the cleavage of benzoyl-
arginine p-nitroanilide (BAPNA) as an assay for peptidase activity. We
purified an arginyl peptidase 168-fold, with a yield of 37%. SDS-PAGE of
the purified fraction revealed a major band of 79 kDa and a fainter one of 41
kDa. N-terminal amino-acid sequencing identified the smaller protein as
aldolase, while the N-terminus of the larger protein was blocked. Activity
assay identified the larger protein as the active peptidase. Fractionation of
the peptidase on a sizing column suggested a size of 188 kDa for the native
protein. Only Arg-containing peptides were cleaved by the peptidase, and it
was found most sensitive to the inhibitors 3,4-dichloroisocoumarin and
antipain. This peptidase could not cleave stromal or thylakoid proteins,
suggesting that its activity is limited to later stages of the protein
degradation
process in chloroplasts, in which short peptides are further cleaved to
ultimately yield free amino acids.
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