CHARACTERIZATION OF THE ALGINATE BIOSYNTHETIC GENE-CLUSTER IN PSEUDOMONAS-SYRINGAE PV SYRINGAE

Alginate, a copolymer of D-mannuronic acid and L-guluronic acid, is pr oduced bg a variety of pseudomamads, including Pseudomonas syringae. A lginate biosynthesis has been most extensively studied in P. aeruginos a, and a number of structural and regulatory genes from this species h ave been cloned and characterized. In the present study, an alginate-d efective (ALg(-)) mutant of P. syringae pv. syringae FF5 was shown to contain a Tn5 insertion in algL, a gene encoding alginate lyase. A cos mid clone designated pSK2 restored alginate production to the algL mut ant and was shown to contain homologs of algD, alg8, alg44 algG, algX (alg60), algL, algF, and algA. The order and arrangement of the struct ural gene cluster were virtually identical to those previously describ ed for P. aeruginosa. Complementation analyses, however, indicated tha t the structural gene clusters in P. aeruginosa and P. syringae were n ot functionally interchangeable when expressed from their native promo ters. A region upstream of the algD gene in P. syringae pv. syringae w as shown to activate the transcription of a promoterless glucuronidase (uidA) gene and indicated that transcription initiated upstream of al gD as described for, P. aeruginosa. Transcription of the algD promoter from P. syringae FF5 was significantly higher at 32 degrees C than at 18 or 26 degrees C and was stimulated when copper sulfate or sodium c hloride was added to the medium. Alginate gene expression was also sti mulated by the addition of the nonionic solute sorbitol, indicating th at osmolarity is a signal for algD expression in P. syringae FF5.