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KMID : 0613820140240020128
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Journal of Life Science
2014 Volume.24 No. 2 p.128 ~ p.136
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Homology Modeling and Active Sites of PolyMG-specific Alginate Lyase from Stenotrophomonas maltophilia KJ-2
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Kim Hee-Sook
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Abstract
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Alginates are linear acidic polysaccharides composed with (1-4)-linked α-L-guluronic acid and β-Dmannuronic acid. Alginate can be degraded by diverse alginate lyases, which cleave the alginate using a β-elimination reaction and produce unsaturated uronate oligomers. A gene for a polyMG-specific alginate lyase possessing a novel structure was previously identified and cloned from Stenotrophomonas maltophilia KJ-2. Homology modeling of KJ-2 polyMG-specific alginate lyase showed it belongs to the PL6 family, whereas three Azotobacter vinelandii polyMG lyases belong to the PL7 family of polysaccharide lyases. From ¹H-NMR spectra data, KJ-2 polyMG lyase preferably degraded the M-β(1-4)-G glycosidic bond than the G-α(1-4)-M glycosidic bond. Seventeen mutants were made by site-directed mutagenesis, and alginate lyase activity was analyzed. Lys220Ala, Arg241Ala, Arg241Lys, and Arg265Ala lost alginate lyase activity completely. Arg155Ala, Gly303Glu, and Tyr304Phe also lost the activity by 60.7-80.1%. These results show that Arg155, Lys220, Arg241, Arg265, Gly303, and Tyr304 are important residues for catalytic activity and substrate binding.
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KEYWORD
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Alginate, alginate lyase, polyMG-specific alginate lyase, homology modeling, site-directed mutagenesis
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