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 cx-amylase (PPA), usin g maltopentaose as the substrate. Lineweaver-Burk plots showed that th e inhibitory action of acarbose is of the mixed non-competitive type. The secondary plots gave straight lines. A model involving abortive co mplexes accounts for these results. Dixon plot analysis led to the sam e conclusion. According to the proposed model, one molecule of acarbos e per amylase molecule binds either directly to free enzyme at the act ive site or to the enzyme-substrate complex at a secondary carbohydrat e-binding site, which becomes functional after the substrate has bound to the enzyme molecule at the active site. Kinetic analysis of the in hibition exerted by either the maltose or maltotriose reaction product s of maltopentaose hydrolysis were then performed. The inhibitory effe ct 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 i s maltose, and that the second product is maltotriose. This indicates that after hydrolysis of the maltopentaose chain, the reducing side fr agment is released first.