Phosphorylation of lipopolysaccharides in the Antarctic psychrotroph Pseudomonas syringae: a possible role in temperature adaptation

Abstract

Phosphorylation of lipopolysaccharide (LPS) from a psychrotrophic bacterium, Pseudomonas syringae, from Antarctica was studied by using sucrose gradient-separated membrane fractions. The bacterium was found to possess an LPS kinase which could phosphorylate more LPS postsynthetically at higher temperatures. The phosphorylation was low at a lower temperature and was also found to occur in vivo. After phosphorylation of LPS in vitro, it was found that the major part of the radioactivity (>85%) was associated with the core oligosaccharide region of the LPS. The phosphate groups of this region are probably involved in the binding of metal ions, which could be removed by EDTA. The cells grown at the lower temperature probably contained fewer divalent cations because of the smaller amount of phosphate and thereby were more sensitive to EDTA. The cells were also more sensitive to cationic antibiotics at the lower temperature. A possible role of this differential phosphorylation of LPS in modulating the function of the outer membrane as a permeability barrier in the psychrotroph is discussed.The outermost layer of the outer membrane bilayer of gram-negative bacteria is composed of lipopolysaccharides (LPS) which contain a hydrophobic moiety, lipid A, and a polysaccharide moiety consisting of the so-called core oligosaccharide and 0 polysaccharide. The chemical structures of lipid A from various bacteria such as Escherichia coli, Pseudomonas aeruginosa, Proteus mirabilis, Salmonella minnesota, Salmonella typhimurium, Shigella sonnei, Yersinia pestis, Er-winia carotovora, and Haemophilus ducreyi are remarkably identical in that they have a backbone of 3-1',6-glucosamine (GlcNAc) (19,32). Heterogeneity of lipid A arises because of variation in the fatty acid composition, which appears to vary depending on the bacterial strain and growth conditions (6,11,21,25,39,40,41,49,52). The composition of the core oligosaccharide is also variable; this oligosaccharide is generally composed of 2-keto-3-deoxyoctonic acid, heptose, glucose, and galactose. The structure of the 0 polysaccharide is most variable, and the 0 polysaccharide may contain various unusual sugars (32,38). The other striking feature of LPS is the presence of a high level of phosphorus (2 to 5.6%), which is attached either to the core sugars (such as 2-keto-3-deoxyoctonic acid and heptose) or to GlcNAc of lipid A as ethanolamine phosphate, ethanolamine PPi, PP1, or simply phosphate (13,25,38,51).Although much information about LPS composition and structure has accumulated during the last 25 years, very little is known about the physiological function of LPS. LPS have been studied as toxins and virulence factors of gram-negative bacteria that cause pathogenesis in animals and plants (19,32). However, the function of LPS with respect to the physiology of the bacteria is poorly understood. In bacteria, LPS has been implicated in providing a kind of permeation barrier for hydrophobic compounds to move across the membrane. The evidence for this has come from the study of...

Influence of Growth Temperature on Lipid and Phosphate Contents of Surface Polysaccharides from the Antarctic Bacterium Pseudoalteromonas haloplanktis TAC 125

“…A quantitative analysis performed on 4°C and 15°C LOS samples revealed that in both cases the phosphate content was lower than that of the 25°C sample by a factor of 4. This finding is in agreement with the suggested increase in LPS kinase activity at higher temperatures, as found for the Antarctic psychrotroph Pseudomonas syringae (16). More recently, in the case of a gram-positive psychrophilic bacterium which lacks LPSs, the amount of phosphoglycerol was found to be greater at 24°C than at 4°C (13).…”

A Unique Capsular Polysaccharide Structure from the Psychrophilic Marine Bacterium Colwellia psychrerythraea 34H That Mimics Antifreeze (Glyco)proteins

“…[1][2][3] Because microorganisms are at thermal equilibrium with their environment, it is reasonable to assume that structural and functional components in psychrophiles (optimal growth at ≤ 15°C) have adapted, to some degree, to the requirements of a low temperature existence, 4 including the possible presence of ice crystals in their immediate surroundings. The reported mechanisms of bacterial adaptation to low temperature include the over-expression of cold-shock and heat-shock proteins, the presence of unsaturated and branched fatty acids that maintain membrane fluidity, 5 the different phosphorylation of membrane proteins and lipopolysaccharides, [6][7][8][9][10][11] and the production of cold-active enzymes, 12 antifreeze proteins and cryoprotectants. 13 The latter are chemical substances that generally include small molecules, such as glycine betaine, some amino acids, sugars (glucose, fructose) and sugar alcohols (mannitol, glycerol).…”

Low-Temperature-Induced Changes in Composition and Fluidity of Lipopolysaccharides in the Antarctic Psychrotrophic BacteriumPseudomonas syringae

“…The LPS from low (4°C)-and high (22°C)-temperature-grown cells of P. syringae were isolated by aqueous phenol (35) as well as by the modified phenol-chloroform-light petroleum method of Brade and Galanos (1). The protease, DNase, and RNase digestions of LPS and the final purification by ultracentrifugation were carried out as described earlier (21). The two procedures (1, 35) yielded similar results showing both S-and R-type LPS (Fig.…”

“…The relationship of the presence of hydroxylated fatty acid residues and the fluidity remains a topic for further study. It was shown earlier that the same strain of Antarctic P. syringae produced, at a lower growth temperature, LPS with lower phosphate content in the core region (21). The possible interaction between the lower fluidity and the altered phosphate FIG.…”

“…Although similar investigations on the change in OM at a low temperature have been conducted with some mesophilic bacteria (11,16,22,23,31,36), such studies on cold-adapted bacteria do not appear to exist. For this reason, we have initiated some studies with the Antarctic psychrotrophic bacterium Pseudomonas syringae, which has the ability to grow at 0 to 4°C (20,21,26). In this report we examine the alteration in OM property and related compositional change in the LPS of low (4°C)-and high (22°C)-temperature-grown cells of P. syringae and correlate them with the fluidity of the hydrophobic core of LPS aggregates in vitro.…”