Exploring amino acids responsible for the temperature profile of glycoside hydrolase family 45 endoglucanase EGL3 from Humicola grisea.
EGL3 and RCE1 are glycoside hydrolase family 45 endoglucanases isolated from Humicola grisea and Rhizopus oryzae respectively. The amino acid sequences of the two endoglucanases are homologous; on the other hand, the optimum temperature of EGL3 is higher than that of RCE1. In this study, four chimeric endoglucanases, named ER1, ER2, ER3 and ER4, in which one of four sequential amino acid regions of the EGL3 catalytic domain (CAD) was replaced by the corresponding RCE1 amino acids, were constructed to explore the region responsible for the EGL3 temperature profile. Then their temperature profiles were compared with that of the recombinant EGL3. Replacement of the N-terminal region of EGL3 with that of RCE1 caused the EGL3 temperature profile to shift to a lower temperature. These results suggest that the N-terminal amino acids of the EGL3 are responsible for the EGL3 temperature profile.
Murashima
Koichiro
K
Food and Health R & D Laboratories, Meiji Seika Kaisha, Ltd., Sakado-shi, Saitama, Japan. koichiro_murashima@meiji.co.jp
Shimonaka
Atsushi
A
Nishimura
Tomoko
T
Baba
Yuko
Y
Koga
Jinichiro
J
Kubota
Hidetoshi
H
Kono
Toshiaki
T
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001203:Ascomycota / Q000201:enzymology* / Q000235:genetics; D002266:Carboxymethylcellulose Sodium / Q000378:metabolism; D002480:Cellulase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050506:Mutant Chimeric Proteins / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D016415:Sequence Alignment; D013329:Structure-Activity Relationship; D013696:Temperature
D000596:Amino Acids; D004271:DNA, Fungal; D050506:Mutant Chimeric Proteins; D002480:Cellulase; D002266:Carboxymethylcellulose Sodium