Conclusions In this study, we were able to clarify the roles of the seven flagellin subunits in the assembly of the flagellar filament in R. leguminosarum. Taken altogether, our results indicate that FlaA is an essential subunit, but that it is not enough to assemble a fully functional flagellar filament. FlaB and FlaC are major components
of the filament while FlaD, FlaE, FlaH, and FlaG are only minor components. To assemble a fully functional filament, at least three (FlaA, FlaB, and FlaC) and five (FlaA, FlaB, FlaC, FlaE, and FlaG) flagellin subunits should be synthesized by 3841 and VF39SM, respectively. There were GDC-0994 manufacturer no substantial differences in the requirements for individual flagellins MI-503 ic50 in swimming vs. swarming motility. The flagellins of 3841 and VF39SM are possibly modified by glycosylation. Acknowledgements We gratefully acknowledge the support for this work from Natural Sciences and Engineering Research Council of Canada (NSERC) Discovery Grants to MFH and SFK. DDT was supported by a Government of Canada graduate scholarship and the Bettina Bahlsen scholarship. We thank Carol Stremick for her help with the protein work as well as Wei-Xiang Dong at the Microscopy and Imaging Facility
of the University of Calgary for his assistance with electron microscopy. We also thank Dr. Christopher K. Yost for his very helpful comments on the manuscript. Electronic supplementary material Additional file 1: Sequences of primers used to PCR amplify flagellin genes. Table showing PCR primer sequences for all PCR work discussed in the paper. (DOC 38 KB) Additional file 2: Details of flagellin gene mutations in R. leguminosarum strains 3841 and VF39SM. Table giving complete description of fragments and cassettes used in construction
of all the mutants described in the paper. (DOC 48 KB) Additional Resveratrol file 3: Immunoblot using an anti-flagellar antibody against flagellar preparations of R. leguminosarum. Figure showing western blot of flagellar preparations of wild type and mutant strains. (PDF 101 KB) Additional file 4: MS/MS spectrum of one tryptic peptide from the data set for VF39SM. Figure showing a Mass Spectrum of a peptide from the tryptic digest of VF39SM flagellar proteins. (PDF 47 KB) References 1. Silverman M: Building bacterial flagella. Q Rev Biol 1980,55(4):395–408.PubMedCrossRef 2. Macnab RM: How bacteria assemble flagella. Annu Rev Microbiol 2003,57(1):77–100.PubMedCrossRef 3. Enomoto M, Sakai A, Tominaga A: Expression of an Escherichia coli flagellin gene, hag48 , in the presence of a Salmonella H1-repressor. Mol Gen Genet 1985,201(1):133–135.PubMedCrossRef 4. Kuwajima G, Asaka J, Fujiwara T, Node K, Kondo E: Nucleotide sequence of the hag gene encoding flagellin of Escherichia coli . J Bacteriol 1986,168(3):1479–1483.PubMed 5.