Biochemical Characterization and Low-Resolution SAXS Molecular Envelope of GH1 β-Glycosidase from Saccharophagus degradans.
The marine bacteria Saccharophagus degradans (also known as Microbulbifer degradans), are rod-shaped and gram-negative motile γ-proteobacteria, capable of both degrading a variety of complex polysaccharides and fermenting monosaccharides into ethanol. In order to obtain insights into structure-function relationships of the enzymes, involved in these biochemical processes, we characterized a S. degradans β-glycosidase from glycoside hydrolase family 1 (SdBgl1B). SdBgl1B has the optimum pH of 6.0 and a melting temperature Tmof approximately 50 °C. The enzyme has high specificity toward short D-glucose saccharides with β-linkages with the following preferences β-1,3 > β-1,4 ≫ β-1,6. The enzyme kinetic parameters, obtained using artificial substrates p-β-NPGlu and p-β-NPFuc and also the disaccharides cellobiose, gentiobiose and laminaribiose, revealed SdBgl1B preference for p-β-NPGlu and laminaribiose, which indicates its affinity for glucose and also preference for β-1,3 linkages. To better understand structural basis of the enzyme activity its 3D model was built and analysed. The 3D model fits well into the experimentally retrieved low-resolution SAXS-based envelope of the enzyme, confirming monomeric state of SdBgl1B in solution.
Instituto de Física de São Carlos, Universidade de São Paulo, Avenida Trabalhador São Carlense 400, São Carlos, SP, 13566-590, Brazil.
Hevila
H
Brognaro
Instituto de Química, Universidade de São Paulo, Avenida Prof. Lineu Prestes, 748, Bloco 10, Sala 1054, São Paulo, SP, 05508-900, Brazil.
Vitor Medeiros
VM
Almeida
Instituto de Física de São Carlos, Universidade de São Paulo, Avenida Trabalhador São Carlense 400, São Carlos, SP, 13566-590, Brazil.
Evandro Ares
EA
de Araujo
Instituto de Física de São Carlos, Universidade de São Paulo, Avenida Trabalhador São Carlense 400, São Carlos, SP, 13566-590, Brazil.
Vasily
V
Piyadov
Instituto de Física de São Carlos, Universidade de São Paulo, Avenida Trabalhador São Carlense 400, São Carlos, SP, 13566-590, Brazil.
Maria Auxiliadora Morim
MA
Santos
Instituto de Química, Universidade de São Paulo, Avenida Prof. Lineu Prestes, 748, Bloco 10, Sala 1054, São Paulo, SP, 05508-900, Brazil.
Sandro Roberto
SR
Marana
Instituto de Física de São Carlos, Universidade de São Paulo, Avenida Trabalhador São Carlense 400, São Carlos, SP, 13566-590, Brazil. ipolikarpov@ifsc.usp.br.
Igor
I
Polikarpov
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020134:Catalytic Domain; D020563:Gammaproteobacteria / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D005959:Glucosidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D053838:Scattering, Small Angle; D013379:Substrate Specificity; D013395:Sucrose / Q000378:metabolism*; D044366:Transition Temperature; D014961:X-Ray Diffraction
D001426:Bacterial Proteins; D013395:Sucrose; D005959:Glucosidases
Enzymatic characterization; Glycosidase hydrolase family 1; Modelling; SAXS structure; Saccharophagus degradans
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