title: Construction of individual, fused, and co-expressed proteins of endoglucanase and β-glucosidase for hydrolyzing sugarcane bagasse.
abstract: At least a combination of endoglucanase (EglII) and β-glucosidase (BglZ) is required for hydrolyzing crystalline cellulose. To understand the catalytic efficiency of combination enzymes for converting biomass to sugars, EglII and BglZ were constructed in the form of individual, fused as well as co-expression proteins, and their activities for hydrolyzing sugarcane bagasse were evaluated. The genes, eglII isolated from Bacillus amyloliquefaciens PSM3.1 earlier and bglZ from B. amyloliquefaciens ABBD, were expressed extracellularly in Bacillus megaterium MS941. EglII exhibited both exoglucanase and endoglucanase activities, and BglZ belonging to the glycoside hydrolase 1 family (GH 1) showed β-glucosidase activity. A combination of EglII and BglZ showed activity on substrates Avicel, CMC and sugarcane bagasse. Specifically for hydrolyzing sugarcane bagasse, fused protein (fus-EglII+BglZ), co-expression protein (coex-BglZ+EglII), and mixed-individual protein (mix-EglII+BglZ) produced cellobiose as the main product, along with a small amount of glucose. The amount of reducing sugars released from the hydrolyzing bleached sugarcane bagasse (BSB) using fus-EglII+BglZ and mix-EglII+BglZ was 2.7- and 4.2-fold higher, respectively, than steamed sugarcane bagasse (SSB), indicating the synergetic enzymes worked better on treated sugarcane bagasse. Compared with fus-EglII+BglZ and mix-EglII+BglZ, coex-BglZ+EglII released more mol reducing sugars from SSB, indicating the enzymes were potential for biomass conversion. Additionally, coex-BglZ+EglII acted on BSB 2.5-fold faster than fus-EglII+BglZ. Thus, coex-bglZ+eglII expression system was the best choice to produce enzymes for hydrolyzing sugarcane baggase.
authors.affiliation: Biochemistry Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesha 10, Bandung 40132, Indonesia.
authors.forename: Sari Dewi
authors.initials: SD
authors.lastname: Kurniasih
authors.affiliation: Biochemistry Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesha 10, Bandung 40132, Indonesia.
authors.forename: Almasul
authors.initials: A
authors.lastname: Alfi
authors.affiliation: Biochemistry Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesha 10, Bandung 40132, Indonesia.
authors.forename: Dessy
authors.initials: D
authors.lastname: Natalia
authors.affiliation: Research Center for Tropical Marine Biotechnology, Diponegoro University, Jl. Soedarto SH No. 1, Semarang 50275, Indonesia.
authors.forename: Ocky Karna
authors.initials: OK
authors.lastname: Radjasa
authors.affiliation: Biochemistry Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesha 10, Bandung 40132, Indonesia. Electronic address: zeily@chem.itb.ac.id.
authors.forename: Zeily
authors.initials: Z
authors.lastname: Nurachman
mesh_terms: D001407:Bacillus / Q000201:enzymology* / Q000235:genetics*; D002266:Carboxymethylcellulose Sodium / Q000378:metabolism; D002480:Cellulase / Q000235:genetics / Q000378:metabolism*; D002482:Cellulose / Q000378:metabolism*; D015870:Gene Expression; D006868:Hydrolysis; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D031786:Saccharum / Q000737:chemistry*; D001617:beta-Glucosidase / Q000235:genetics / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D002482:Cellulose; C027433:bagasse; D001617:beta-Glucosidase; D002480:Cellulase; D002266:Carboxymethylcellulose Sodium
keywords: Co-expressed protein; Endoglucanase; Fused protein; Sugarcane bagasse; β-Glucosidase