Binding of the substrate UDP-glucuronic acid induces conformational changes in the xanthan gum glucuronosyltransferase

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-tri...

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Detalles Bibliográficos
Autores principales: Salinas, Silvina R., Petruk, Ariel Alcides, Brukman, Nicolás G., Martí, Marcelo Adrián, Ielpi, Luis
Publicado: 2016
Materias:
Conformational change
Glycosyltransferase
GumK
Membrane monotopic protein
Xanthan
Bins
Biochemistry
Circular dichroism spectroscopy
Conformations
Dichroism
Glucose
Molecular dynamics
Proteins
Conformational differences
Glycosyl transferase
Molecular dynamics simulations
Substrate-binding sites
Xanthomonas campestris
Xanthan gum
glucuronosyltransferase
unclassified drug
uridine diphosphate glucuronic acid
xanthan
xanthan gum glucuronosyltransferase
bacterial polysaccharide
protein aggregate
protein binding
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
protein conformation
Binding Sites
Glucuronosyltransferase
Molecular Docking Simulation
Molecular Dynamics Simulation
Polysaccharides, Bacterial
Protein Aggregates
Protein Binding
Protein Conformation
Uridine Diphosphate Glucuronic Acid
Acceso en línea:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_17410126_v29_n6_p197_Salinas
http://hdl.handle.net/20.500.12110/paper_17410126_v29_n6_p197_Salinas
Aporte de:
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) de Universidad de Buenos Aires
Descripción
Sumario: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.

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