Structural characterization of the Acetobacter xylinum endo-beta-1,4-glucanase CMCax required for cellulose biosynthesis.
Previous studies have demonstrated that endoglucanase is required for cellulose biosynthesis both in bacteria and plants. However, it has yet to be elucidated how the endoglucanases function in the mechanism of cellulose biosynthesis. Here we describe the crystal structure of the cellulose biosynthesis-related endo-beta-1,47-glucanase (CMCax; EC 3.2.1.4) from the cellulose-producing Gramnegative bacterium, Acetobacter xylinum (= Gluconacetobacter xylinus), determined at 1.65-A resolution. CMCax falls into the glycoside hydrolase family 8 (GH-8), and the structure showed that the overall fold of the CMCax is similar to those of other glycoside hydrolases belonging to GH-8. Structure comparison with Clostridium thermocellum CelA, the best characterized GH-8 endoglucanase, revealed that sugar recognition subsite +3 is completely missing in CMCax. The absence of the subsite +3 leads to significant broadness of the cleft at the cellooligosaccharide reducing-end side. CMCax is known to be a secreted enzyme and is present in the culture medium. However, electron microscopic analysis using immunostaining clearly demonstrated that a portion of CMCax is localized to the cell surface, suggesting a link with other known membrane-anchored endoglucanases that are required for cellulose biosynthesis.
Yasutake
Yoshiaki
Y
Division of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo, Hokkaido 060-0810, Japan.
Kawano
Shin
S
Tajima
Kenji
K
Yao
Min
M
Satoh
Yasuharu
Y
Munekata
Masanobu
M
Tanaka
Isao
I
D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000201:enzymology; D002480:Cellulase / Q000737:chemistry* / Q000378:metabolism; D002482:Cellulose / Q000096:biosynthesis*; D003460:Crystallization; D018360:Crystallography, X-Ray; D041822:Gluconacetobacter xylinus / Q000201:enzymology* / Q000648:ultrastructure; D046529:Microscopy, Electron, Transmission; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000378:metabolism; D016415:Sequence Alignment
D009844:Oligosaccharides; D002482:Cellulose; D002480:Cellulase