D-Xylose detection in Escherichia coli by a xylose binding protein-dependent response

Author(s)

Ribeiro, Lucas F. ^[1] ; Bressan, Fabiana ^[2] ; Furtado, Gilvan P. ^[1] ; Meireles, Flavio ^[2] ; Ward, Richard J. ^{[3, 4]}

Affiliation

^[1] Univ Sao Paulo, Fac Med Ribeirao Preto, Dept Bioquim & Imunol, BR-14049900 Ribeirao Preto, SP - Brazil

^[2] Fac Zootecnia & Engn Alimentos, Dept Med Vet, Pirassununga, SP - Brazil

^[3] Univ Sao Paulo, Fac Filosofia Ciencias & Letras Ribeirao Preto, Dept Quim, BR-14049901 Ribeirao Preto, SP - Brazil

^[4] CNPEM, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, Sao Paulo - Brazil

• Document type: Journal article
• Source: Journal of Biotechnology; v. 168, n. 4, p. 440-445, DEC 2013.
• Web of Science Citations: 3

Abstract

A gene circuit for the controlled expression of a marker gene and for the assay of xylose concentration in Escherichia coli has been designed and tested. The xylF coding sequence for the xylose binding protein (XBP) was cloned in pT7T318U downstream from the promoter for xylanase A from B. subtilis (Pbsu), together with the GFP coding sequence (gfp) under the control of the xylF promoter, forming the pT7T3-GFP-XBP construct. GFP fluorescence in Escherichia coli JW3538-1 xylF-transformed with pT7T3-GFP-XBP was approximately 1.4x higher after 520 min growth in the presence of 5 mM xylose than in cells transformed with pT7T3-GFP. Under saturating xylose concentration, flow cytometry analysis showed that all cells resulted in homogeneous populations, and the population with XBP showed a fluorescence greater than that without XBP. Activity of the xylF promoter in cells transformed with pT7T3-GFP-XBP was similar to 40% higher than with the pT7T3-GFP. No response was observed with arabinose and ribose, showing that the expression effects were specific for xylose, demonstrating the potential use of the gene circuit as a biosensor. (C) 2013 Elsevier B.V. All rights reserved. (AU)

• FAPESP's process: 10/18850-2 - Identification, characterization and engineering of plant cell wall degrading enzymes
• Grantee: Richard John Ward
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 10/07133-8 - Creation of an Allosteric Endoxylanase by Directed Evolution for application in Industrial Biotechnology
• Grantee: Lucas Ferreira Ribeiro
• Support Opportunities: Scholarships in Brazil - Doctorate