The mechanism of porcine pancreatic alpha-amylase. Inhibition of maltopentaose hydrolysis by acarbose, maltose and maltotriose.
A kinetic study was carried out on the inhibitory effects of acarbose, maltose, and maltotriose on porcine pancreatic alpha-amylase (PPA), using maltopentaose as the substrate. Lineweaver-Burk plots showed that the inhibitory action of acarbose is of the mixed non-competitive type. The secondary plots gave straight lines. A model involving abortive complexes accounts for these results. Dixon plot analysis led to the same conclusion. According to the proposed model, one molecule of acarbose per amylase molecule binds either directly to free enzyme at the active site or to the enzyme-substrate complex at a secondary carbohydrate-binding site, which becomes functional after the substrate has bound to the enzyme molecule at the active site. Kinetic analysis of the inhibition exerted by either the maltose or maltotriose reaction products of maltopentaose hydrolysis were then performed. The inhibitory effect of maltose was found to be of the non-competitive type, while that of maltotriose was competitive. It can therefore be concluded that the first reaction product to be released upon maltopentaose hydrolysis is maltose, and that the second product is maltotriose. This indicates that after hydrolysis of the maltopentaose chain, the reducing side fragment is released first.
Al Kazaz
M
M
Laboratoire de Biochimie et Biologie de la Nutrition CNRS-UPRESA 6033, Faculté des Sciences et Techniques de St Jérôme, Université d'Aix-Marseille, Marseille, France.
Desseaux
V
V
Marchis-Mouren
G
G
Prodanov
E
E
Santimone
M
M
D020909:Acarbose; D000818:Animals; D001665:Binding Sites / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007700:Kinetics; D008320:Maltose / Q000494:pharmacology; D009844:Oligosaccharides / Q000378:metabolism; D010179:Pancreas / Q000201:enzymology*; D011485:Protein Binding / Q000502:physiology; D013552:Swine; D014312:Trisaccharides / Q000494:pharmacology; D000516:alpha-Amylases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D004791:Enzyme Inhibitors; D009844:Oligosaccharides; D014312:Trisaccharides; C019193:maltopentaose; C008317:maltotriose; D008320:Maltose; D000516:alpha-Amylases; D020909:Acarbose