Structural and antigenic characteristics of Campylobacter coli FlaA flagellin.
The polar flagellar filament of Campylobacter coli VC167 is composed of two highly related (98%) flagellin subunit proteins, FlaA and FlaB, whose antigenic specificities result from posttranslational modification. FlaA is the predominant flagellin species, and mutants expressing only FlaA form a full-length flagellar filament. Although the deduced M(r) of type 2 (T2) FlaA is 58,884 and the apparent M(r) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis is 59,500, the solution weight-average M(r) by sedimentation analysis was 63,000. Circular dichroism studies in the presence or absence of 0.1% sodium dodecyl sulfate or 50% trifluorethanol showed that the secondary structure of T2 FlaA flagellin was altered, with alpha-helix structure being increased to 25% in the nonpolar environment. The molecule also contained 35 to 48% beta-sheet and 11 to 29% beta-turn structure. Mimeotope analysis of octapeptides representing the sequence of FlaA together with immunoelectron microscopy and enzyme-linked immunosorbent assay with a panel of antisera indicated that many residues in presumed linear epitopes were inaccessible or nonepitopic in the assembled filament, with the majority being in the N-terminal 337 residues of the 572-residue flagellin. Residues at the carboxy-terminal end of the T2 FlaA subunit also become inaccessible upon assembly. Digestion with trypsin, chymotrypsin, and endoproteinase Glu-C revealed a protease-resistant domain with an approximate M(r) of 18,700 between residues 193 and 375. Digestion with endoproteinase Arg-C and endoproteinase Lys-C allowed the mapping of a segment of surface-exposed FlaA sequence which contributes serospecificity to the VC167 T2 flagellar filament at residues between 421 and 480.
Power
M E
ME
Department of Biochemistry and Microbiology, University of Victoria, British Columbia, Canada.
Guerry
P
P
McCubbin
W D
WD
Kay
C M
CM
Trust
T J
TJ
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D017000:Campylobacter coli / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D002942:Circular Dichroism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D005407:Flagella / Q000737:chemistry / Q000276:immunology*; D005408:Flagellin / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D008666:Metalloendopeptidases / Q000378:metabolism; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017433:Protein Structure, Secondary; D012697:Serine Endopeptidases / Q000378:metabolism; D012703:Serotyping; D013056:Spectrophotometry, Ultraviolet
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D000939:Epitopes; D005408:Flagellin; C067138:flaA protein, bacteria; C044499:submandibular proteinase A; D012697:Serine Endopeptidases; D008666:Metalloendopeptidases; C039830:peptidyl-Lys metalloendopeptidase