title: Multifunctional cellulolytic auxiliary activity protein HcAA10-2 from Hahella chejuensis enhances enzymatic hydrolysis of crystalline cellulose.
abstract: The modular auxiliary activity (AA) family of proteins is believed to cause amorphogenesis in addition to oxidative cleavage of crystalline cellulose although the supporting evidence is limited. HcAA10-2 is a modular AA10 family protein (58 kDa) composed of a AA10 module and a family two carbohydrate binding module (CBM2), joined by a long stretch of 222 amino acids of unknown function. The protein was expressed in Escherichia coli and purified to homogeneity. Scanning electron microscopy and X-ray diffraction analysis of Avicel treated with HcAA10-2 provided evidence for the disruption of the cellulose microfibrils ("amorphogenesis") and reduction of the crystallinity index, resulting in a twofold increase of cellulase adsorption on the polysaccharide surface. HcAA10-2 exhibited weak endoglucanase-like activity toward soluble cellulose and cello-oligosaccharides with an optimum at pH 6.5 and 45 °C. HcAA10-2 catalyzed oxidative cleavage of crystalline cellulose released native and oxidized cello-oligosaccharides in the presence of copper and an electron donor such as ascorbic acid. Multiple sequence alignment indicated that His1, His109, and Phe197 in the AA10 module formed the conserved copper-binding site. The reducing sugar released from Avicel by the endoglucanase Cel5 and Celluclast accompanying HcAA10-2 was increased by four- and sixfold, respectively. Moreover, HcAA10-2 and Celluclast acted synergistically on pretreated wheat straw biomass resulting in a threefold increase in reducing sugar than Celluclast alone. Taken together, these results suggest that HcAA10-2 is a novel multifunctional modular AA10 protein possessing amorphogenesis, weak endoglucanase, and oxidative cleavage activities useful for efficient degradation of crystalline cellulose.
authors.affiliation: Division of Applied Life Sciences (BK21), SSAC and PMBBRC, Gyeongsang National University, Jinju, 660-701, Republic of Korea.
authors.forename: Sunil S
authors.initials: SS
authors.lastname: Ghatge
authors.forename: Amar A
authors.initials: AA
authors.lastname: Telke
authors.forename: Tatoba R
authors.initials: TR
authors.lastname: Waghmode
authors.forename: Yuno
authors.initials: Y
authors.lastname: Lee
authors.forename: Keun-Woo
authors.initials: KW
authors.lastname: Lee
authors.forename: Doo-Byoung
authors.initials: DB
authors.lastname: Oh
authors.forename: Hyun-Dong
authors.initials: HD
authors.lastname: Shin
authors.forename: Seon-Won
authors.initials: SW
authors.lastname: Kim
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D001709:Biotechnology / Q000379:methods*; D002480:Cellulase / Q000737:chemistry / Q000378:metabolism; D002482:Cellulose / Q000737:chemistry / Q000378:metabolism*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D020563:Gammaproteobacteria / Q000737:chemistry* / Q000235:genetics; D006868:Hydrolysis; D008670:Metals / Q000378:metabolism; D008855:Microscopy, Electron, Scanning; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014908:Triticum; D014961:X-Ray Diffraction
chemical_list: D001426:Bacterial Proteins; D008670:Metals; D011994:Recombinant Proteins; D002482:Cellulose; C407024:endoglucanase Cel5A; D002480:Cellulase