2Institute of Cell Biophysics, Pushchino Scientific Center for Biological Research, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia
* To whom correspondence should be addressed.
Received April 18, 2022; Revised May 30, 2022; Accepted May 30, 2022
Cellulophaga lytica is a Gram-negative aerobic bacterium in the genome of which there are many genes encoding polysaccharide degrading enzymes. One of the enzymes named ClGP contains a glycoside hydrolase domain from the GH5 family and a polysaccharide lyase domain from the PL31 family. The enzyme also contains the TAT signaling peptide and the TIGR04183 domain that indicates extracellular nature of the enzyme. Phylogenetic analysis has shown that the enzymes most closely related to ClGP and containing all four domains (TAT, GH5, PL31, TIGR04183) are widespread among bacterial species belonging to the Flavobacteriaceae family. ClGP produced by the recombinant strain of E. coli was purified and characterized. ClGP exhibited activity of endoglucanase (EC 22.214.171.124) and catalyzed hydrolysis of β-D-glucan, carboxymethyl cellulose sodium salt (CMC-Na), and amorphous cellulose, but failed to hydrolyze microcrystalline cellulose and xylan. Products of CMC hydrolysis were cellobiose and cellotriose, whereas β-D-glucan was hydrolyzed to glucose, cellobiose, cellotetraose, and cellopentaose. ClGP was more active against the poly-β-D-mannuronate blocks than against the poly-α-L-glucuronate blocks of alginic acid. This indicates that the enzyme is a polyM lyase (EC 126.96.36.199). ClGP was active against polyglucuronic acid, so it displayed a glucuronan lyase (EC 188.8.131.52) activity. The enzyme had a neutral pH-optimum, was stable in the pH range 6.0-8.0, and displayed moderate thermal stability. ClGP effectively saccharified two species of brown algae, Saccharina latissima and Laminaria digitata, that suggests its potential for use in the production of biofuel from macroalgae.
KEY WORDS: GH5 endoglucanase, PL31 alginase, multifunctional enzyme, saccharification of macroalgae