Studies on the active site of human alpha-amylases: examination of the third subsite S3' of the aglycone-binding site by control of substrate binding mode.
Modified maltooligosaccharides, IG-G-G-G-AG-G (IG: 6-deoxy-6-iodo-D-glucopyranose residue, G: D-glucopyranose residue, AG: 6-amino-6-deoxy-D-glucopyranose residue, -: alpha-1,4-glycosidic linkage), IG-G-G-G-AG-M (M: methyl), and IG-G-G-G-AG-phi (phi: phenyl) were prepared by the use of cyclodextrin glucanotransferase in order to examine the third subsite (S3') of the aglycone-binding site of human salivary and pancreatic alpha-amylases. Human alpha-amylases hydrolyzed the modified maltooligosaccharides to IG-G-G and G-AG-G, IG-G-G and G-AG-M, and IG-G-G and G-AG-phi. This implied that G, M, and phi fit into S3'. There was no difference in the rate parameters between the two enzymes. The Km values for the hydrolysis of IG-G-G-G-AG-G by both enzymes were the same as those for IG-G-G-G-AG-M, and twice those for IG-G-G-G-AG-phi. The results showed that S3' of the two enzymes has no affinity for the glucose residue and is not a subsite but a hydrophobic environment.
Omichi
K
K
Department of Chemistry, Osaka University College of Science.
Hase
S
S
Ikenaka
T
T
D001407:Bacillus / Q000201:enzymology; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000378:metabolism*; D010189:Pancreatic Juice / Q000201:enzymology; D012469:Salivary Glands / Q000201:enzymology; D016339:Spectrometry, Mass, Fast Atom Bombardment; D000516:alpha-Amylases / Q000737:chemistry* / Q000378:metabolism
D009844:Oligosaccharides; D000516:alpha-Amylases