Blackwell PublishingActa Medica Okayama1462-5814862006Identification of glycosylation genes and glycosylated amino acids of flagellin in Pseudomonas syringae pv. tabaci923938ENFumikoTaguchiKasumiTakeuchiEtsukoKatohKatsuyoshiMurataTomokoSuzukiMizuriMarutaniTakayukiKawasakiMinakoEguchiShizueKatohHanaekakuChihiroYasudaYoshishigeInagakiKazuhiroToyodaTomonoriShiraishiYukiIchinoseA glycosylation island is a genetic region required for glycosylation. The glycosylation island of flagellin in Pseudomonas syringae pv. tabaci 6605 consists of three orfs: orf1, orf2 and orf3. Orf1 and orf2 encode putative glycosyltransferases, and their deletion mutants, Delta orf1 and Delta orf2, exhibit deficient flagellin glycosylation or produce partially glycosylated flagellin respectively. Digestion of glycosylated flagellin from wild-type bacteria and non-glycosylated flagellin from Delta orf1 mutant using aspartic N-peptidase and subsequent HPLC analysis revealed candidate glycosylated amino acids. By generation of site-directed Ser/Ala-substituted mutants, all glycosylated amino acid residues were identified at positions 143, 164, 176, 183, 193 and 201. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry (MS) analysis revealed that each glycan was about 540 Da. While all glycosylation-defective mutants retained swimming ability, swarming ability was reduced in the Delta orf1, Delta orf2 and Ser/Ala-substituted mutants. All glycosylation mutants were also found to be impaired in the ability to adhere to a polystyrene surface and in the ability to cause disease in tobacco. Based on the predicted tertiary structure of flagellin, S176 and S183 are expected to be located on most external surface of the flagellum. Thus the effect of Ala-substitution of these serines is stronger than that of other serines. These results suggest that glycosylation of flagellin in P. syringae pv. tabaci 6605 is required for bacterial virulence. It is also possible that glycosylation of flagellin may mask elicitor function of flagellin molecule.No potential conflict of interest relevant to this article was reported.Blackwell PublishingActa Medica Okayama0108-27016312007Hydrogen bonding in two solid phases of phenazine-chloranilic acid (1/1) determined at 170 and 93 Ko17o20ENKazumaGotohTetsuoAsajiHiroyukiIshidaThe crystal structures in two solid phases, i.e. phase II stable between 146 and 253 K and phase IV below 136 K, of the title compound [phenazine-chloranilic acid (1/1), C12H8N2 center dot C6H2Cl2O4, in phase II, and phenazinium hydrogen chloranilate, C12H9N2+center dot C6HCl2O4-, in phase IV], have been determined. Both phases crystallize in P2(1), and each structure was refined as an inversion twin. In phase II, the phenazine and chloranilic acid molecules are arranged alternately through two kinds of O-H center dot center dot center dot N hydrogen bonds. In phase IV, salt formation occurs by donation of one H atom from the chloranilic acid molecule to the phenazine molecule; the resulting monocation and monoanion are linked by N-H center dot center dot center dot O and O-H center dot center dot center dot N hydrogen bonds.No potential conflict of interest relevant to this article was reported.