Molecular identification of Sphingomonas sp. A1 alginate lyase (A1-IV') as a member of novel polysaccharide lyase family 15 and implications in alginate lyase evolution.
Sphingomonas sp. A1 (strain A1) produces three endotypes (A1-I [65 kDa], A1-II [25 kDa], and A1-III [40 kDa]) and an exotype (A1-IV [86 kDa]) alginate lyases in cytoplasm. These four enzymes cooperatively depolymerize alginate into constituent monosaccharides. In addition to the genes for these lyases, novel genes encoding hypothetical proteins homologous with A1-IV were found in the genomes of many bacteria including strain A1. One such protein, A1-IV' (90 kDa) of strain A1, was overexpressed in Escherichia coli cells, purified, and characterized. A1-IV' catalyzed the cleavage of glycosidic bonds in alginate through a beta-elimination reaction and released unsaturated di- and trisaccharides as main products, thus indicating that the enzyme is an endotype alginate lyase. A1-IV', which differed from A1-IV in some enzymatic properties, was not expressed in strain A1, suggesting that A1-IV' has no significant role in alginate metabolism. A1-IV' and other A1-IV homologs facilitate the creation of novel polysaccharide lyase family 15 based on their primary structures, implying the evolution route of alginate lyases in family PL-15.
Hashimoto
Wataru
W
Laboratory of Basic and Applied Molecular Biotechnology, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
Miyake
Osamu
O
Ochiai
Akihito
A
Murata
Kousaku
K
D000595:Amino Acid Sequence; D017124:Conserved Sequence; D019143:Evolution, Molecular; D008969:Molecular Sequence Data; D011133:Polysaccharide-Lyases / Q000737:chemistry* / Q000235:genetics*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D020578:Sphingomonas / Q000201:enzymology* / Q000235:genetics*
D011994:Recombinant Proteins; D011133:Polysaccharide-Lyases; C016158:poly(beta-D-mannuronate) lyase