title: Crystal structure and genetic modifications of FI-CMCase from Aspergillus aculeatus F-50.
abstract: Cellulose is the major component of the plant cell wall and the most abundant renewable biomass on earth, and its decomposition has proven to be very useful in many commercial applications. Endo-1,4-β-d-glucanase (EC 3.2.1.4; endoglucanase), which catalyzes the random hydrolysis of 1,4-β-glycosidic bonds of the cellulose main chain to cleave cellulose into smaller fragments, is the key cellulolytic enzyme. An endoglucanase isolated from Aspergillus aculeatus F-50 (FI-CMCase), which is classified into the glycoside hydrolase (GH) family 12, was demonstrated to be effectively expressed in the industrial strain Pichia pastoris. Here, the crystal structure and complex structures of P. pastoris-expressed FI-CMCase were solved to high resolution. The overall structure is analyzed and compared to other GH12 members. In addition, the substrate-surrounding residues were engineered to search for variants with improved enzymatic activity. Among 14 mutants constructed, one with two-fold increase in protein expression was identified, which possesses a potential to be further developed as a commercial enzyme product.
authors.affiliation: AsiaPac Biotechnology Co., Ltd, Dongguan 523808, China.
authors.forename: Jian-Wen
authors.initials: JW
authors.lastname: Huang
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Weidong
authors.initials: W
authors.lastname: Liu
authors.affiliation: AsiaPac Biotechnology Co., Ltd, Dongguan 523808, China.
authors.forename: Hui-Lin
authors.initials: HL
authors.lastname: Lai
authors.affiliation: AsiaPac Biotechnology Co., Ltd, Dongguan 523808, China.
authors.forename: Ya-Shan
authors.initials: YS
authors.lastname: Cheng
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Yingying
authors.initials: Y
authors.lastname: Zheng
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Li
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Hong
authors.initials: H
authors.lastname: Sun
authors.affiliation: Department of Veterinary Medicine, National Chung Hsing University, Taichung City, 402, Taiwan, ROC.
authors.forename: Chih-Jung
authors.initials: CJ
authors.lastname: Kuo
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China. Electronic address: guo_rt@tib.cas.cn.
authors.forename: Rey-Ting
authors.initials: RT
authors.lastname: Guo
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China. Electronic address: ccckate0722@gmail.com.
authors.forename: Chun-Chi
authors.initials: CC
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D001228:Aspergillosis / Q000382:microbiology; D001230:Aspergillus / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D002480:Cellulase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray; D015870:Gene Expression; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D010843:Pichia / Q000235:genetics; D011487:Protein Conformation; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D006026:Glycoside Hydrolases; D002480:Cellulase
keywords: Aspergillus aculeatus; Endoglucanase; Protein structure; X-ray crystallography