title: Arginine is essential for the alpha-amylase inhibitory activity of the alpha-amylase/subtilisin inhibitor (BASI) from barley seeds.
abstract: Treatment of barley alpha-amylase/subtilisin inhibitor (BASI) with reagents specific for arginine, histidine, methionine and tyrosine residues and amino and carboxyl groups indicates that an arginine residue(s) is essential for its action on the target enzyme barley alpha-amylase 2. Phenylglyoxal modified eight out of 12 arginine residues in BASI. Kinetic analysis shows that the inactivation of BASI follows a pseudo-first-order reaction and is due to reaction with one molecule of phenylglyoxal; the second-order rate constant is determined to be 2.95 M-1.min-1. At pH 8.0, BASI and barley alpha-amylase 2 form an inactive 1:1 complex. The Ki value of this association is 2.2 x 10(-10) M. The alpha-amylase protects four arginine residues and also the alpha-amylase inhibitory activity of BASI against phenylglyoxal. When BASI from the phenylglyoxal-modified target enzyme-inhibitor complex is isolated and subjected to a second treatment with phenylglyoxal, four additional arginine residues are modified, with concomitant loss of the inhibitory activity. These results are discussed in relation to a three-dimensional model of BASI based on the known structure of the corresponding inhibitor from wheat.
authors.affiliation: Department of Chemistry, Carlsberg Laboratory, Copenhagen Valby, Denmark.
authors.forename: J
authors.initials: J
authors.lastname: Abe
authors.forename: U
authors.initials: U
authors.lastname: Sidenius
authors.forename: B
authors.initials: B
authors.lastname: Svensson
mesh_terms: D001120:Arginine / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002850:Chromatography, Gel; D001467:Hordeum / Q000201:enzymology; D007700:Kinetics; D010658:Phenylglyoxal / Q000494:pharmacology*; D013381:Subtilisins / Q000037:antagonists & inhibitors*; D000516:alpha-Amylases / Q000037:antagonists & inhibitors*
chemical_list: D001426:Bacterial Proteins; C018150:subtilisin inhibitor protein, Streptomyces; D001120:Arginine; D000516:alpha-Amylases; D013381:Subtilisins; D010658:Phenylglyoxal