title: Effects of growth conditions on archaellation and N-glycosylation in Methanococcus maripaludis.
abstract: In this study, the effects of growth conditions on archaellation in Methanococcus maripaludis were examined. Cells were grown in a variety of media, including complex, minimal and with formate as the electron donor, with different nitrogen sources, varied salinities and at a variety of growth temperatures. Of the conditions tested, Western blot results showed that major archaellin FlaB2 levels only varied detectably as a result of growth temperature. Whilst the amount of FlaB2 was similar for cells grown at < 35 °C, protein levels decreased at 38 °C and were barely detectable at 42 °C. Quantitative reverse transcription PCR experiments demonstrated that the flaB2 transcript levels were almost undetectable at 42 °C. Electron microscopy confirmed that the FlaB2 levels detected by Western blots corresponded to the state of archaellation, with cells grown at 42 °C being mostly non-archaellated. Unexpectedly, a lower apparent molecular mass for FlaB2 was observed in Western blots of cells grown at temperatures >38 °C, suggestive of a truncation in the attached N-linked tetrasaccharide at higher growth temperatures. MS analysis of archaella isolated from cells grown at 40 °C confirmed that FlaB2 was now decorated with a trisaccharide in which the third sugar was also lacking the attached threonine and acetamidino modifications found in the WT glycan.
authors.affiliation: Department of Biomedical and Molecular Sciences,Queen's University, Kingston, Ontario K7L 3N6,Canada.
authors.forename: Yan
authors.initials: Y
authors.lastname: Ding
authors.affiliation: Department of Biomedical and Molecular Sciences,Queen's University, Kingston, Ontario K7L 3N6,Canada.
authors.forename: Zoe
authors.initials: Z
authors.lastname: Lau
authors.affiliation: Human Health Therapeutics Portfolio,National Research Council, Ottawa, Ontario K1A 0R6,Canada.
authors.forename: Susan M
authors.initials: SM
authors.lastname: Logan
authors.affiliation: Human Health Therapeutics Portfolio,National Research Council, Ottawa, Ontario K1A 0R6,Canada.
authors.forename: John F
authors.initials: JF
authors.lastname: Kelly
authors.affiliation: Department of Molecular and Cellular Biology,University of Guelph, Guelph, Ontario N1G 2W1,Canada.
authors.forename: Alison
authors.initials: A
authors.lastname: Berezuk
authors.affiliation: Department of Molecular and Cellular Biology,University of Guelph, Guelph, Ontario N1G 2W1,Canada.
authors.forename: Cezar M
authors.initials: CM
authors.lastname: Khursigara
authors.affiliation: Department of Biomedical and Molecular Sciences,Queen's University, Kingston, Ontario K7L 3N6,Canada.
authors.forename: Ken F
authors.initials: KF
authors.lastname: Jarrell
mesh_terms: D019843:Archaeal Proteins / Q000235:genetics / Q000378:metabolism*; D005408:Flagellin / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D006358:Hot Temperature; D017017:Methanococcus / Q000254:growth & development* / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D011134:Polysaccharides / Q000378:metabolism*
chemical_list: D019843:Archaeal Proteins; C489684:FlaB2 protein, archaeal; D011134:Polysaccharides; D005408:Flagellin