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Molecular cloning and expression of a new α‐neoagarobiose hydrolase from Agarivorans gilvus WH0801 and enzymatic production of 3,6‐anhydro‐ l ‐galactose
Author(s) -
Liu Nan,
Yang Meng,
Mao Xiangzhao,
Mu Bozhong,
Wei Dongzhi
Publication year - 2015
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1002/bab.1363
Subject(s) - hydrolase , biochemistry , chemistry , molecular mass , enzyme , agarose , hydrolysis , cloning (programming) , galactose , molecular cloning , amino acid , glycoside hydrolase , recombinant dna , affinity chromatography , peptide sequence , microbiology and biotechnology , gene , biology , computer science , programming language
A new α‐neoagarobiose hydrolase (NABH) called AgaWH117 was cloned from Agarivorans gilvus WH0801. The gene encoding this hydrolase consists of 1,086 bp and encodes a protein containing 361 amino acids. This new NABH showed 74% amino acid sequence identity with other known NABHs. The molecular mass of the recombinant AgaWH117 was estimated to be 41 kDa. Purified AgaWH117 showed endolytic activity during neoagarobiose degradation, yielding 3,6‐anhydro‐ l ‐galactose ( l ‐AHG) and d ‐galactose as products. It showed a maximum activity at a temperature of 30 °C and a pH of 6.0 and was stable at temperatures below 30 °C. Its K m and V max values were 2.094 mg/mL and 6.982 U/mg, respectively. The cloning strategy used and AgaWH117 isolated in this study will provide information on the saccharification process of marine biomass. This study provides a method to produce l ‐AHG from agarose by using AgaWH117 without an acid and describes its one‐step purification by using Bio‐Gel P2 chromatography.