Functional analysis of the Xylella fastidiosa gum operon and its relation to the citrus variegated clorosis (CVC)

• Grant number: 98/16260-0
• Support Opportunities: Genome Research Grants
• Start date: March 01, 1999
• End date: December 31, 2001
• Field of knowledge: Biological Sciences - Biochemistry
• Principal Investigator: Paulo Arruda
• Grantee: Paulo Arruda
• Host Institution: Instituto de Biologia (IB). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil

Abstract

Sequence analysis of a -12 Kb DNA fragment of the 7 A2 cosmid of the Xylella fastidiosa Genome Project, revealed the presence of an operon containing 9 genes (B, C, D, E, F, H, J, K e M) homologous to the Xanthomonas campestris Gum operon. The X. campestris Gum operon encodes the enzymes responsible for the biosynthesis of the exopolysaccharide xanthan gum, involved in pathogenicity. Deletion of the gumD gene of aX. campestris strain reduced virulence (Chou et al, BBRC, 1997, 233: 265-269) and alterations in the later stages of xanthan biosynthesis reduced the bacteria aggressiveness against plant (Katzen et ai, 1998, J Bacteriol. 180: 1607-1617). Thus it is possible that the X. fastidiosa when introduced into the xylem of orange plants produces an exopolisaccharide wich occlude the vessels leading to the variegated clorosis. In this project we shall determine if the X. fastidiosa Gum operon is functional. The exopolysaccharide produced will be chemically characterized and its presence in infected plants will be examined. Key genes of the biosynthetic pathway will be expressed in E. coli and the enzymes characterized. The product of the gumD gene will be purified to homogeneity and the 3-0 structure of the protein will be determined. This will be used for further investigation of a possible disease control system. The production of extracellular enzymes such as protease, amylase, cellulase and pectinase, which is involved with pathogenicity in other bacteria, will be investigated. (AU)