Isolation and partial characterization of the extracellular polysaccharides and lipopolysaccharides from fast-growing Rhizobium japonicum USDA 205 and its Nod- mutant, HC205, which lacks the symbiotic plasmid

Abstract

The extracellular polysaccharides and lipopolysaccharides (LPSs) from two fast-growing Rhizobium japonicum strains, USDA 205 and HC205, were isolated and partially characterized. Strain HC205 is a Nod- mutant of USDA 205 which lacks the symbiotic plasmid. The extracellular polysaccharides from both strains are very similar in composition, having galactose, glucose, glucuronic acid, and acyl groups. The extracellular polysaccharides do not contain detectable levels of pyruvate. Methylation analysis shows that the extracellular polysaccharides from both strains have the same glycosyl linkages. The LPSs were purified by a modified phenol-water extraction procedure and gel filtration chromatography. The LPSs from USDA 205 and HC205 elute as broad peaks from the gel filtration column and contain 2-keto-3-deoxyoctonic acid as one of the major sugar components. Each broad 2-keto-3-deoxyoctonic acid-containing peak has a distinct shoulder on its leading edge. The shoulder and the remainder of the broad peak are separated and labeled LPSI and LPSII, respectively. Glucose (and 2-keto-3-deoxyoctonic acid) is a major sugar in the LPSI fractions. Both the LPSII fractions contain 2-keto-3-deoxyoctonic acid as the major sugar (about 20% of the mass). There are a number of quantitative differences in these LPS fractions between strain USDA 205 and HC205. Polyacrylamide gel electrophoresis shows that the LPSs are heterogeneous molecules but less heterogeneous than the LPSs from Salmonella minnesota or Rhizobium leguminosarum. The LPSI fractions from both USDA 205 and HC205 show a single lower-molecular-weight band and a higher-molecular-weight banding region which contains several bands. No bands are observed for the LPSII fractions from either USDA 205 or HC205.