Open AccessA New Acrosin Inhibitor from Boar Spermatozoa
Author(s) -
TSCHESCHE Harald,
WITTIG Bernd,
DECKER Günther,
MÜLLERESTERL Werner,
FRITZ Hans
Publication year - 1982
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1982.tb06752.x
Subject(s) - acrosin , chemistry , sephadex , biochemistry , isoelectric point , trypsin , size exclusion chromatography , kunitz sti protease inhibitor , trypsin inhibitor , affinity chromatography , chromatography , enzyme , biology , acrosome , sperm , botany
A new proteinase inhibitor ( M r 7500) was isolated to apparent homogeneity from boar spermatozoa by repeated gel filtration on Sephadex G‐50 and affinity chromatography on concanavalin‐A‐Sepharose 4B. The inhibitor strongly inhibits boar acrosin in a competitive 1:1 stoichiometric reaction with a K ass = 7 × 10 10 l mol −1 . The inhibitor is a glycoprotein and represents a first member of a new class of proteinase inhibitor with a rather short polypeptide backbone of only 42 amino acid residues and a low cystine content. The basic protein (isoelectric point 9.4) contains a single disulfide loop, which is easily reducible by sodium borohydride. Upon reduction the inhibitory activity is lost, but rapidly regained after air reoxidation of the corresponding half‐cystine residues. The reactive site residue was established to be arginine by inhibition with 2,3‐butanedione. The inhibitor is rather specific for acrosin and inhibits bovine trypsin only to a limited extent. However, incubation with catalytic amounts of trypsin (or acrosin) at acid pH (pH 2–3) rapidly leads to a limited proteolysis at the reactive site with formation of 67% modified (reactive site hydrolysed), but still active inhibitor. The equilibrium constant was established to be K hyd = 2.0.