Hydrolysis of ionic liquid–treated substrate with an Iocasia fonsfrigidae strain SP3-1 endoglucanase

• Published in: Applied microbiology and biotechnology Vol. 108; no. 1; p. 63
• Main Authors: Heng, Sobroney, Sutheeworapong, Sawannee, Wangnai, Chinnapong, Champreda, Verawat, Kosugi, Akihiko, Ratanakhanokchai, Khanok, Tachaapaikoon, Chakrit, Ceballos, Ruben Michael
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer Berlin Heidelberg 01.12.2024; Springer Nature B.V
• Subjects: bacteria; Bacteria, Anaerobic; barley; Base Composition; biomass; Biomedical and Life Sciences; Biotechnologically Relevant Enzymes and Proteins; Biotechnology; Carboxymethyl cellulose; Carboxymethylcellulose; Cellobiose; Cellulase; Cellulolytic enzymes; Cellulose; Chromatography; Crystalline cellulose; endo-1,4-beta-glucanase; Endoglucanase; enzymatic hydrolysis; Enzymes; family; feedstocks; Glucan; Glucans; Glycosidases; Glycoside hydrolase; glycosides; High performance liquid chromatography; Hydrolysis; hypersalinity; Ionic Liquids; Kinetics; Life Sciences; Liquid chromatography; mechanism of action; Microbial Genetics and Genomics; Microbiology; Mode of action; Phylogeny; RNA, Ribosomal, 16S; Saline solutions; Salinity tolerance; Salt tolerance; Sequence Analysis, DNA; sodium; Sodium Chloride; Substrate specificity; Substrates; Thailand; Thin layer chromatography; Thailand; Ionic liquids; Endoglucanase; Salt tolerance; strain SP3-1; Glycoside hydrolase family 12; Enzyme platform; Iocasia fonsfrigidae strain SP3-1
• ISSN: 0175-7598; 1432-0614; 1432-0614
• DOI: 10.1007/s00253-023-12918-1

Recently, we reported the discovery of a novel endoglucanase of the glycoside hydrolase family 12 (GH12), designated IfCelS12A, from the haloalkaliphilic anaerobic bacterium Iocasia fonsfrigidae strain SP3-1, which was isolated from a hypersaline pond in the Samut Sakhon province of Thailand (ca. 2017). IfCelS12A exhibits high substrate specificity on carboxymethyl cellulose and amorphous cellulose but low substrate specificity on b-1,3;1,4-glucan. Unlike some endoglucanases of the GH12 family, IfCelS12A does not exhibit hydrolytic activity on crystalline cellulose (i.e., Avicel™). High-Pressure Liquid Chromatography (HPLC) and Thin Layer Chromatography (TLC) analyses of products resulting from IfCelS12-mediated hydrolysis indicate mode of action for this enzyme. Notably, IfCelS12A preferentially hydrolyzes cellotetraoses, cellopentaoses, and cellohexaoses with negligible activity on cellobiose or cellotriose. Kinetic analysis with cellopentaose and barely b- d -glucan as cellulosic substrates were conducted. On cellopentaose, IfCelS12A demonstrates a 16-fold increase in activity ( K M  = 0.27 mM; k cat  = 0.36 s −1 ; k cat /K M  = 1.34 mM −1 s −1 ) compared to the enzymatic hydrolysis of barley b- d -glucan ( K M : 0.04 mM, k cat : 0.51 s −1 , k cat /K M  = 0.08 mM −1 s −1 ). Moreover, IfCelS12A enzymatic efficacy is stable in hypersaline sodium chlorids (NaCl) solutions (up to 10% NaCl). Specifically, IfCel12A retains notable activity after 24 h at 2M NaCl (10% saline solution). IfCelS12A used as a cocktail component with other cellulolytic enzymes and in conjunction with mobile sequestration platform technology offers additional options for deconstruction of ionic liquid–pretreated cellulosic feedstock. Key points • IfCelS12A from an anaerobic alkaliphile Iocasia fronsfrigidae shows salt tolerance • IfCelS12A in cocktails with other enzymes efficiently degrades cellulosic biomass • IfCelS12A used with mobile enzyme sequestration platforms enhances hydrolysis