A bifunctional endolytic alginate lyase with two different lyase catalytic domains from Vibrio sp. H204
Chune Peng, Qingbin Wang, Wei Xu, Xinkun Wang, Qianqian Zheng, Xiaohui Liang, Xiaodan Dong, Fuchuan Li, Lizeng Peng
Abstract
Alginate lyases can fully degrade alginate into various size-defined unsaturated oligosaccharide products by β -elimination. Here, we identified the bifunctional endolytic alginate lyase Aly35 from the marine bacterium Vibrio sp. Strain H204. The enzyme Aly35 is classified into the polysaccharide lyase 7 superfamily and contains two alginate lyase catalytic domains. The relationship and function of the two lyase domains are not well known. Thus, the full-length recombinant enzyme and its truncated proteins Aly35-CD1 (catalytic domain 1), Aly35-CD2 (catalytic domain 2 domain) were constructed. The three enzymes showed similar biochemical characteristics and exhibited temperature and pH stability. Further research showed that Aly35 and Aly35-CD2 can efficiently degrade alginate, polymannuronate (PM) and polyguluronate (PG) into a series of unsaturated oligosaccharides, while Aly35-CD1 exhibits greater PM-degrading activity than that of Aly35-CD2 but can not degraded PG efficiently. The results suggest that the domain (Trp 295 -His 582 ) is critical for PG-degrading activity, the domain has (Leu 53 -Lys 286 ) higher PM-degrading activity, both catalytic domains together confer increased alginate (including M-blocks and G blocks)-degrading activity. The enzyme Aly35 and its truncations Aly35-CD1 and Aly35-CD2 will be useful tools for structural analyses and for preparing bioactive oligosaccharides, especially Aly35-CD1 can be used to prepare G unit–rich oligosaccharides from alginate.