title: Enzymatic properties and primary structures of two α-amylase isozymes from the Pacific abalone Haliotis discus hannai.
abstract: Two α-amylase (EC 3.2.1.1) isozymes, HdAmy58 and HdAmy82, with approximate molecular masses of 58 kDa and 82 kDa, respectively, were isolated from the digestive fluid of the Pacific abalone Haliotis discus hannai. Optimal temperatures and pHs for HdAmy58 and HdAmy82 were at 30 °C and 6.7, and 30 °C and 6.1, respectively. Both enzymes similarly degraded starch, glycogen, and maltooligosaccharides larger than maltotriose producing maltose and maltotriose as the major degradation products. However, the activity toward maltotetraose was appreciably higher in HdAmy82 than HdAmy58. cDNAs encoding HdAmy58 and HdAmy82 were cloned and the amino-acid sequences of 511 and 694 residues for HdAmy58 and HdAmy82, respectively, were deduced. The putative catalytic domains of HdAmy58 and HdAmy82 were located in the 17-511th and 19-500th amino-acid regions, respectively, and they showed approximately 50% amino-acid identity to each other. These sequences also showed 62-99% amino-acid identity to the catalytic domains of known α-amylases that belong to glycoside-hydrolase-family 13. The difference in the molecular masses between HdAmy58 and HdAmy82 was ascribed to the extension of approximately 190 residues in the C-terminus of HdAmy82. This extended region showed 41-63% amino-acid identity with the ancillary domains of several α-amylases previously reported.
authors.affiliation: Laboratory of Marine Biotechnology and Microbiology, Graduate School of Fisheries Sciences, Hokkaido University, Minato-cho 3-1-1, Hakodate 041-8611, Japan.
authors.forename: Yuya
authors.initials: Y
authors.lastname: Kumagai
authors.forename: Takuya
authors.initials: T
authors.lastname: Satoh
authors.forename: Akira
authors.initials: A
authors.lastname: Inoue
authors.forename: Takao
authors.initials: T
authors.lastname: Ojima
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D055162:Biocatalysis; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D049851:Gastropoda / Q000201:enzymology* / Q000235:genetics*; D006003:Glycogen / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009844:Oligosaccharides / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013213:Starch / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D000516:alpha-Amylases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D018076:DNA, Complementary; D007527:Isoenzymes; D009844:Oligosaccharides; C021705:maltooligosaccharides; D013213:Starch; D006003:Glycogen; D000516:alpha-Amylases