Advanced search
Start date
(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Real-time investigation of mannosyltransferase function of a Xylella fastidiosa recombinant GumH protein using QCM-D

Full text
Alves, Claudia A. [1] ; Pedroso, Mariele M. [1] ; de Moraes, Marcela C. [1] ; Souza, Dulce H. F. [1] ; Cass, Quezia B. [1] ; Faria, Ronaldo C. [1]
Total Authors: 6
[1] Univ Fed Sao Carlos, Dept Quim, BR-13565905 Sao Carlos, SP - Brazil
Total Affiliations: 1
Document type: Journal article
Source: Biochemical and Biophysical Research Communications; v. 408, n. 4, p. 571-575, MAY 20 2011.
Web of Science Citations: 2

Xylella fastidiosa is a gram-negative bacterium that causes serious diseases in economically important crops, including grapevine, coffee, and citrus fruits. X. fastidiosa colonizes the xylem vessels of the infected plants, thereby blocking water and nutrient transport. The genome sequence of X. fastidiosa has revealed an operon containing nine genes possibly involved in the synthesis of an exopolisaccharide (EPS) named fastidian gum that can be related with the pathogenicity of this bacterium. The alpha-1,3-mannosyltransferase (GumH) enzyme from X. fastidiosa is involved in fastidian gum production. GumH is responsible for the transfer of mannose from guanosine diphosphate mannose (GDP-man) to the cellobiose-pyrophosphate-polyprenol carrier lipid (CPP-Lip) during the assembly and biosynthesis of EPS. In this work, a method for real-time detection of recombinant GumH enzymatic activity was successfully developed using a Quartz Crystal Microbalance with dissipation monitoring (QCM-D). The QCM-D transducer was strategically modified with CPP-Lip by using a solid-supported lipid bilayer that makes use of a self-assembled monolayer of 1-undecanethiol. Monitoring the real-time CPP-Lip QCM-D transducer in the presence of GDP-man and GumH enzyme shows a mass increase, indicating the transfer of mannose. The real-time QCM-D determination of mannosyltransferase function was validated by a High Performance Liquid Chromatography (LC) method developed for determination of GDP produced by enzymatic reaction. LC results confirmed the activity of recombinant GumH protein, which is the first enzyme involved in the biosynthesis of the EPS from X. fastidiosa enzymatically characterized. (C) 2011 Elsevier Inc. All rights reserved. (AU)

FAPESP's process: 08/06867-8 - Development, characterization and analytical applications of nanocomposites electrodes in catecholamines and flavonoids determination
Grantee:Ronaldo Censi Faria
Support type: Regular Research Grants