Cel8H, a novel endoglucanase from the halophilic bacterium Halomonas sp. S66-4: molecular cloning, heterogonous expression, and biochemical characterization.
A recombinant Escherichia coli clone expressing an endoglucanase was identified from a genomic library of the halophilic bacterium Halomonas sp. S66-4, and the enzyme was designated Cel8H. The cel8H gene consisted of 1,053 bp and encoded 350 amino acids sharing the highest identity of 48% to other known endoglucanases. The protein was expressed in E. coli BL21 (DE3) and purified to homogeneity. The purified recombinant enzyme had an optimal activity of 4.9 U/mg at pH 5 and 45 degrees C toward the substrate carboxymethylcellulose. It exhibited extraordinary properties which differed from endoglucanases reported previously at the point of high salt tolerance above 5 M, simultaneously with high pH stability at pH 4-12 and high temperature stability at 40-60 degrees C. Various substrate tests indicated that the enzyme hydrolyzes beta-1,4-glucosidic bonds specifically.
Huang
Xiaoluo
X
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, PR China.
Shao
Zongze
Z
Hong
Yuzhi
Y
Lin
Ling
L
Li
Chanjuan
C
Huang
Fei
F
Wang
Hui
H
Liu
Ziduo
Z
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D002266:Carboxymethylcellulose Sodium / Q000378:metabolism; D002480:Cellulase / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D015698:Genomic Library; D020632:Halomonas / Q000201:enzymology* / Q000235:genetics*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013379:Substrate Specificity; D013696:Temperature
D001426:Bacterial Proteins; D017931:DNA Primers; D004269:DNA, Bacterial; D011994:Recombinant Proteins; D002480:Cellulase; D002266:Carboxymethylcellulose Sodium