Evidence for essential arginine residues at the active sites of maize branching enzymes.
Alignment of 23 branching enzyme (BE) amino acid sequences from various species showed conservation of two arginine residues. Phenylglyoxal (PGO) was used to investigate the involvement of arginine residues of maize BEI and BEII in catalysis. BE was significantly inactivated by PGO in triethanolamine buffer at pH 8.5. The inactivation followed a time- and concentration-dependent manner and showed pseudo first-order kinetics. Slopes of 0.73 (BEI) and 1.05 (BEII) were obtained from double log plots of the observed rates of inactivation against the concentrations of PGO, suggesting that loss of BE activity results from as few as one arginine residue modified by PGO. BE inactivation was positively correlated with [14C]PGO incorporation into BE protein and was considerably protected by amylose and/or amylopectin, suggesting that the modified arginine residue may be involved in substrate binding or located near the substrate-binding sites of maize branching enzymes I and II.
Cao
H
H
Department of Biochemistry, Michigan State University, East Lansing 48824, USA.
Preiss
J
J
D015061:1,4-alpha-Glucan Branching Enzyme / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics; D000595:Amino Acid Sequence; D000687:Amylopectin / Q000378:metabolism; D000688:Amylose / Q000378:metabolism; D000818:Animals; D001120:Arginine / Q000737:chemistry*; D001665:Binding Sites; D002417:Cattle; D017124:Conserved Sequence; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000032:analysis / Q000737:chemistry*; D004926:Escherichia coli; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D010658:Phenylglyoxal / Q000032:analysis / Q000737:chemistry*; D010762:Phosphorylase a / Q000378:metabolism*; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011817:Rabbits; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics; D016415:Sequence Alignment; D011198:Solanum tuberosum; D003313:Zea mays / Q000201:enzymology
D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D000688:Amylose; D000687:Amylopectin; D001120:Arginine; D010762:Phosphorylase a; D015061:1,4-alpha-Glucan Branching Enzyme; D010658:Phenylglyoxal