The pH dependence of the action pattern in porcine pancreatic alpha-amylase-catalyzed reaction for maltooligosaccharide substrates.
Porcine pancreatic alpha-amylase (EC 3.2.1.1; abbreviated PPA), which hydrolyzes alpha-D-(1,4) glucosidic bonds in starch and amylose, displays an optimum at pH 6.9 for the majority of substrates. The optimum pH, however, shifted to 5.2 for the hydrolysis of some low molecular substrates (Ishikawa, K., et al., 1990, Biochemistry 29, 7119-7123). Details of the substrate-dependent shift of the optimum pH in PPA were studied by use of a series of maltooligosaccharides with 14C-labeled reducing end glucose as substrates. The optimum pH for maltotriose was 5.2, whereas that for maltopentaose and maltohexaose was unchanged at pH 6.9. The pH profile for the intermediate size substrate maltotetraose showed abnormality; the apparent optimum pH was broadened between 5.5 and 6.5 and the bond cleavage pattern depended on pH, unlike that for the other substrates examined. These results were independent of either buffer systems or substrate concentration. Analyses of the hydrolysates of the maltooligosaccharides revealed that the shift of the optimum pH to the neutral region occurred only when the fifth subsite of PPA in the productive binding modes was occupied by a glucosyl residue of a substrate. The three-catalytic residue model of PPA deduced from the analysis of the hydrolysis of some modified maltooligosaccharides (p-nitrophenyl-alpha-D-maltoside, gamma-cyclodextrin, maltopentaitol, and maltohexaitol) (Ishikawa, K., et al., 1990, Biochemistry 29, 7119-7123) was successfully adapted to the linear maltooligosaccharides used in this work. These results indicate that the different productive binding modes of the linear oligosaccharide substrates affect directly the catalytic power and the optimum pH of PPA.
Ishikawa
K
K
National Chemical Laboratory for Industry, Ibaraki, Japan.
Matsui
I
I
Honda
K
K
Kobayashi
S
S
Nakatani
H
H
D000818:Animals; D001665:Binding Sites; D002021:Buffers; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D009844:Oligosaccharides / Q000378:metabolism*; D010179:Pancreas / Q000201:enzymology*; D013552:Swine; D000516:alpha-Amylases / Q000378:metabolism*
D002021:Buffers; D009844:Oligosaccharides; C021705:maltooligosaccharides; D000516:alpha-Amylases