Overexpression and characterization of an extremely thermostable maltogenic amylase, with an optimal temperature of 100 degrees C, from the hyperthermophilic archaeon Staphylothermus marinus.
A gene encoding a hyperthermostable maltogenic amylase of Staphylothermus marinus (SMMA) was cloned and overexpressed in Escherichia coli. SMMA consisted of 696 amino acids with a predicted molecular mass of 82.5 kDa. The enzyme was active in acidic conditions (pH 3.5-5.0), with an optimal pH of 5.0, and was extremely thermostable, with a temperature optimum of 100 degrees C and a melting temperature of 109 degrees C, both of which extremely favored the starch conversion process. SMMA hydrolyzed linear malto-oligosaccharides, starch, cyclodextrins, and cycloamylose, primarily to maltose and glucose, and showed highest activity toward acarbose and pullulan, hydrolyzed to acarviosine-glucose and panose, respectively. Investigation of the cleavage mode using (14)C-maltoheptaose revealed that SMMA preferentially hydrolyzed the first and second glycosidic bonds from the reducing end. To our knowledge, this enzyme is the most thermostable maltogenic amylase yet reported, and might be of potential value in the food and starch industries.
Li
Dan
D
Center for Agricultural Biomaterials, and Department of Food Science and Biotechnology, School of Agricultural Biotechnology, Seoul National University, Seoul 151-921, Republic of Korea.
Park
Jong-Tae
JT
Li
Xiaolei
X
Kim
Sukyung
S
Lee
Seungjae
S
Shim
Jae-Hoon
JH
Park
Sung-Hoon
SH
Cha
Jaeho
J
Lee
Byong-Hoon
BH
Kim
Jung-Wan
JW
Park
Kwan-Hwa
KH
D000595:Amino Acid Sequence; D002152:Calorimetry, Differential Scanning; D002855:Chromatography, Thin Layer; D019620:Desulfurococcaceae / Q000201:enzymology* / Q000235:genetics; D004795:Enzyme Stability; D019766:Genes, Archaeal / Q000235:genetics; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010802:Phylogeny; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D016415:Sequence Alignment; D013379:Substrate Specificity; D013696:Temperature; D014461:Ultracentrifugation
D006026:Glycoside Hydrolases; C120909:glucan 1,4-alpha-maltohydrolase