Extracellular polysaccharide produced by a virulent strain (U-7) of Pseudomonas solanacearum.

Abstract

An extracellular polysaccharide was isolated from the culture mediumofa virulent strain (U-7) of Pseudomonassolanacearum. The polysaccharide caused tomato cuttings to wilt. It seemed to be composed mainly of N-acetylgalactosamine together with small amounts of rhamnose, glucose, and protein. This is, to our knowledge, the first example of homo-N-acetylgalactosaminoglycan in Packard 3380A digital integrator. GC-MS(70eV) were generated with a Hitachi M-80 mass spectrometer and M-003 data processor. The total sugar content was assayed by the phenol-sulfuric acid method.6) Hexosamine was measured colorimetrically7) after hydrolysis in hydrochloric acid. Sugar was analyzed by GLCof the alditol acetates on a Hewlett-Packard fused-silica capillary column (50 mx0.31 mm).8) The 13C-NMR (125 MHz) spectra for solution of EPS in D2Owere recorded on a Brucker AM-500 spectrometer at 30°C using a 5-mmtube and internal methanol (49.9ppm from Me4Si) as the standard. The infrared (IR) spectrum was obtained using a Nicolet 60SX. Total nitrogen was assayed by elemental analysis. Acetic acid was measured with GLC9) using propionic acid as the internal standard. The GLCconditions were as follows: column, a glass column (100 x 0.3cm) packed with Porapack QS (80~100mesh); carrier gas, N2 at 60ml/min; injection temperature, 250°C; oven temperature, 150°C isothermal. Amino acids were analyzed in a JEOL JLC-6AH amino acid analyzer after hydrolysis of the sample in 6n hydrochloric acid at 110°C for 18hr in a sealed glass

A gene cluster required for coordinated biosynthesis of lipopolysaccharide and extracellular polysaccharide also affects virulence of Pseudomonas solanacearum

“…The sugar rhamnose has been identified in both EPS and LPS of P. solanacearum (1,3). DNA sequencing and comparison with published sequences revealed that opsG encodes two polypeptides which have greater than 70% amino acid identity to the two polypeptides of the rhamnose synthetase of Salmonella typhimurium (22, 26a by pL5001, many other LPS biosynthetic genes apparently exist in P. solanacearum.…”

“…One EPS molecule was reported to be a linear polymer of N-galactosamine (GalNac), with glucose and rhamnose as minor components (1,14). An acidic fraction of EPS was recently purified and shown to contain unusual amino sugars, such as basillosamine, and uronic acids, such as 2-N-acetyl-2-deoxy-Lgalacturonic acid, but not rhamnose (34).…”

Purification and Some Properties of the Endo α-1,4 Polygalactosaminidase from Pseudomonas sp.

“…CM-Sephadex C-25, Sephadex G-50 and Phenyl-Sepharose CL-4B were purchased from Pharmacia Fine Chemicals, Sweden, carrier ampholyte from LKBProdukter AB, Sweden. Polymer and oligomers(n > 6) of N-acetyl a-l, 4 galactosamine were prepared according to the method of Takada et al 9 ) The N-acetyl fJ-l,3 polygalactosamine of Pseudomonas solanacearum 10 ) was kindly supplied by Dr. Akiyama. Nitrophenyl-glycosides and other enzyme substrates were form Seikagaku Kogyo Co., Tokyo.…”

Genetic and biochemical characterization of a Pseudomonas solanacearum gene cluster required for extracellular polysaccharide production and for virulence

“…Most of the known or presumed virulence factors in P. solanacearum are extracellular products, including the plant cell wall-degrading enzymes polygalacturonase (3, 27) and endoglucanase (26) and the plant hormones auxin, cytokinin, and ethylene (9). Although this array of extracellular products would seem to constitute a formidable arsenal for virulence, the most intensively studied virulence factor from P. solanacearum is a relatively inert product, extracellular polysaccharide (EPS) (1,11,13,19,31,36,38). This high-molecular-weight polymer is thought to be responsible for induction of the wilt symptoms that characterize the disease caused by P. solanacearum.…”

“…Akiyama et al (1) suggested that EPS is a homopolymer of N-acetylgalactosamine (GalNAc). Glucose and rhamnose have been reported to be minor components (1,11,13), but the presence of these sugars is probably the result of incomplete purification. Instead, two amino sugars, bacillosamine (2,4-diamino-2,4,6-trideoxyglucose) and galactosaminuronic acid, probably are integral components of the P. solanacearum EPS.…”