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Salinas, S.R.; Petruk, A.A.; Brukman, N.G.; Bianco, M.I.; Jacobs, M.; Marti, M.A.; Ielpi, L. "Binding of the substrate UDP-glucuronic acid induces conformational changes in the xanthan gum glucuronosyltransferase" (2016) Protein Engineering, Design and Selection. 29(6):197-207
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Abstract:

GumK is a membrane-associated glucuronosyltransferase of Xanthomonas campestris that is involved in xanthan gum biosynthesis. GumK belongs to the inverting GT-B superfamily and catalyzes the transfer of a glucuronic acid (GlcA) residue from uridine diphosphate (UDP)-GlcA (UDP-GlcA) to a lipid-PP-trisaccharide embedded in the membrane of the bacteria. The structure of GumK was previously described in its apo- and UDP-bound forms, with no significant conformational differences being observed. Here, we study the behavior of GumK toward its donor substrate UDP-GlcA. Turbidity measurements revealed that the interaction of GumK with UDP-GlcA produces aggregation of protein molecules under specific conditions. Moreover, limited proteolysis assays demonstrated protection of enzymatic digestion when UDP-GlcA is present, and this protection is promoted by substrate binding. Circular dichroism spectroscopy also revealed changes in the GumK tertiary structure after UDP-GlcA addition. According to the obtained emission fluorescence results, we suggest the possibility of exposure of hydrophobic residues upon UDP-GlcA binding. We present in silico-built models of GumK complexed with UDP-GlcA as well as its analogs UDP-glucose and UDP-galacturonic acid. Through molecular dynamics simulations, we also show that a relative movement between the domains appears to be specific and to be triggered by UDP-GlcA. The results presented here strongly suggest that GumK undergoes a conformational change upon donor substrate binding, likely bringing the two Rossmann fold domains closer together and triggering a change in the N-terminal domain, with consequent generation of the acceptor substrate binding site. © 2016 The Author.

Registro:

Documento: Artículo
Título:Binding of the substrate UDP-glucuronic acid induces conformational changes in the xanthan gum glucuronosyltransferase
Autor:Salinas, S.R.; Petruk, A.A.; Brukman, N.G.; Bianco, M.I.; Jacobs, M.; Marti, M.A.; Ielpi, L.
Filiación:Laboratory of Bacterial Genetics, Fundación Instituto Leloir, IIBBA, CONICET, Av. Patricias Argentinas 435, Buenos Aires, C1405BWE, Argentina
Departamento de Química Inorgánica, Analítica, y Química Física, INQUIMAE, CONICET, Córdoba, Argentina
Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires, C1428EHA, Argentina
Departamento de Química Biológica, CIQUIBIC, CONICET, Universidad Nacional de Córdoba, Argentina
Department of Chemistry, Faculty of Science, University of Waterloo, Waterloo, ON, Canada
IBYME, CONICET, Buenos Aires, Argentina
Instituto de Ciencia y Tecnología Dr. César Milstein, Fundación Pablo Cassará, CONICET, Buenos Aires, Argentina
Palabras clave:Conformational change; Glycosyltransferase; GumK; Membrane monotopic protein; Xanthan; Bins; Biochemistry; Circular dichroism spectroscopy; Conformations; Dichroism; Glucose; Molecular dynamics; Proteins; Conformational change; Conformational differences; Glycosyl transferase; GumK; Molecular dynamics simulations; Substrate-binding sites; Xanthan; Xanthomonas campestris; Xanthan gum; glucuronosyltransferase; unclassified drug; uridine diphosphate glucuronic acid; xanthan; xanthan gum glucuronosyltransferase; bacterial polysaccharide; glucuronosyltransferase; protein aggregate; protein binding; uridine diphosphate glucuronic acid; xanthan; amino acid substitution; amino terminal sequence; Article; binding site; carboxy terminal sequence; circular dichroism; complex formation; conformational transition; controlled study; crystal structure; enzyme conformation; enzyme metabolism; enzyme substrate complex; hydrophobicity; ligand binding; membrane binding; molecular docking; molecular dynamics; nonhuman; priority journal; protein aggregation; protein degradation; protein domain; protein protein interaction; turbidity; chemistry; enzymology; metabolism; molecular docking; protein conformation; Xanthomonas campestris; Binding Sites; Glucuronosyltransferase; Molecular Docking Simulation; Molecular Dynamics Simulation; Polysaccharides, Bacterial; Protein Aggregates; Protein Binding; Protein Conformation; Uridine Diphosphate Glucuronic Acid; Xanthomonas campestris
Año:2016
Volumen:29
Número:6
Página de inicio:197
Página de fin:207
DOI: http://dx.doi.org/10.1093/protein/gzw007
Título revista:Protein Engineering, Design and Selection
Título revista abreviado:Protein Eng. Des. Sel.
ISSN:17410126
CODEN:PEDSB
CAS:glucuronosyltransferase, 37329-64-9, 9030-08-4; uridine diphosphate glucuronic acid, 2616-64-0; xanthan, 11138-66-2; Glucuronosyltransferase; Polysaccharides, Bacterial; Protein Aggregates; Uridine Diphosphate Glucuronic Acid; xanthan gum
Registro:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_17410126_v29_n6_p197_Salinas

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Citas:

---------- APA ----------
Salinas, S.R., Petruk, A.A., Brukman, N.G., Bianco, M.I., Jacobs, M., Marti, M.A. & Ielpi, L. (2016) . Binding of the substrate UDP-glucuronic acid induces conformational changes in the xanthan gum glucuronosyltransferase. Protein Engineering, Design and Selection, 29(6), 197-207.
http://dx.doi.org/10.1093/protein/gzw007
---------- CHICAGO ----------
Salinas, S.R., Petruk, A.A., Brukman, N.G., Bianco, M.I., Jacobs, M., Marti, M.A., et al. "Binding of the substrate UDP-glucuronic acid induces conformational changes in the xanthan gum glucuronosyltransferase" . Protein Engineering, Design and Selection 29, no. 6 (2016) : 197-207.
http://dx.doi.org/10.1093/protein/gzw007
---------- MLA ----------
Salinas, S.R., Petruk, A.A., Brukman, N.G., Bianco, M.I., Jacobs, M., Marti, M.A., et al. "Binding of the substrate UDP-glucuronic acid induces conformational changes in the xanthan gum glucuronosyltransferase" . Protein Engineering, Design and Selection, vol. 29, no. 6, 2016, pp. 197-207.
http://dx.doi.org/10.1093/protein/gzw007
---------- VANCOUVER ----------
Salinas, S.R., Petruk, A.A., Brukman, N.G., Bianco, M.I., Jacobs, M., Marti, M.A., et al. Binding of the substrate UDP-glucuronic acid induces conformational changes in the xanthan gum glucuronosyltransferase. Protein Eng. Des. Sel. 2016;29(6):197-207.
http://dx.doi.org/10.1093/protein/gzw007