Purification and characterization of a multienzyme complex produced by Paenibacillus curdlanolyticus B-6.
Paenibacillus curdlanolyticus B-6 showed effective degradation activities for xylan and cellulose and produced an extracellular multienzyme complex (approximately 1,450 kDa) containing several xylanases and cellulases. To characterize the multienzyme complex, we purified the complex from culture supernatants by four kind of chromatography. The purified multienzyme complex was composed of a 280-kDa protein with xylanase activity, a 260-kDa protein that was a truncated form on the C-terminal side of the 280-kDa protein, two xylanases of 40 and 48 kDa, and 60 and 65 kDa proteins having both xylanase and carboxymethyl cellulase activities. The 280-kDa protein resembled the scaffolding proteins of cellulosomes based on its migratory behavior in polyacrylamide gels and as a glycoprotein. Cloning of the 40-kDa major xylanase subunit named Xyn11A revealed that Xyn11A contained two functional domains which belonged to glycosyl hydrolase family-11 and to carbohydrate-binding module family-36, respectively, and a glycine- and asparagine-rich linker. However, an amino acid sequence similar to a dockerin domain, which is crucial to cellulosome assembly, was not found in Xyn11A. These results suggest that the multienzyme complex produced by P. curdlanolyticus B-6 should assemble by a mechanism distinct from the cohesin-dockerin interactions known in cellulosomes.
Pason
Patthra
P
Post-harvest Science and Technology Division, Japan International Research Center for Agricultural Sciences, 1-1 Ohwashi, Tsukuba, Ibaraki, Japan.
Kosugi
Akihiko
A
Waeonukul
Rattiya
R
Tachaapaikoon
Chakrit
C
Ratanakhanokchai
Khanok
K
Arai
Takamitsu
T
Murata
Yoshinori
Y
Nakajima
Jun
J
Mori
Yutaka
Y
D044602:Cellulases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D002845:Chromatography / Q000379:methods; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D056507:Paenibacillus / Q000201:enzymology*; D017434:Protein Structure, Tertiary; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D044602:Cellulases; D043364:Endo-1,4-beta Xylanases