PECTATE LYASE PELI OF ERWINIA-CHRYSANTHEMI-3937 BELONGS TO A NEW FAMILY

Erwinia chrysanthemi 3937 secretes five major isoenzymes of pectate ly ases encoded by the pelA, pelB, pelC, pelD, and pelE genes and a set o f secondary pectate lyases, two of which, pelL and pelZ, have been alr eady identified. We cloned the pelI gene, encoding a ninth pectate lya se of E. chrysanthemi 3937. The pelI reading frame is 1,035 bases long , corresponding to a protein of 344 amino acids including a typical am ino-terminal signal sequence of 19 amino acids. The purified mature Pe lI protein has an isoelectric point of about 9 and an apparent molecul ar mass of 34 kDa. PelI has a preference for partially methyl esterifi ed pectin and presents an endo-cleaving activity with an alkaline pH o ptimum and an absolute requirement for Ca2+ ions. PelI is an extracell ular protein secreted by the Out secretory pathway of E. chrysanthemi. The PelI protein is very active in the maceration of plant tissues. A pelI mutant displayed reduced pathogenicity on chicory leaves, but it s virulence did not appear to be affected on potato tubers or Saintpau lia ionantha plants. The pelI gene constitutes an independent transcri ptional unit. As shown for the other pel genes, the transcription of p elI is dependent on various environmental conditions. It is induced by pectic catabolic products and affected by growth phase, oxygen limita tion, temperature, nitrogen starvation, and catabolite repression. Reg ulation of pelI expression appeared to be dependent on the three repre ssors of pectinase synthesis, KdgR, PecS, and PecT, and on the global activator of sugar catabolism, cyclic AMP receptor protein. A function al KdgR binding site was identified close to the putative pelI promote r. Analysis of the amino acid sequence of PelI revealed high homology with a pectate lyase from Erwinia carotovora subsp. carotovora (65% id entity) and low homology with pectate lyases of the phytopathogenic fu ngus Nectria haematococca (Fusarium solani). This finding indicates th at PelI belongs to pectate lyase class III. Using immunoblotting exper iments, we detected PelI homologs in various strains of E. chrysanthem i and E. carotovora subsp. carotovora but not in E. carotovora subsp. atroseptica.