A lipopolysaccharide-binding domain of the Campylobacter fetus S-layer protein resides within the conserved N terminus of a family of silent and divergent homologs.
Campylobacter fetus cells can produce multiple S-layer proteins ranging from 97 to 149 kDa, with a single form predominating in cultured cells. We have cloned, sequenced, and expressed in Escherichia coli a sapA homolog, sapA2, which encodes a full-length 1,109-amino-acid (112-kDa) S-layer protein. Comparison with the two previously cloned sapA homologs has demonstrated two regions of identity, approximately 70 bp before the open reading frame (ORF) and proceeding 550 bp into the ORF and immediately downstream of the ORF. The entire genome contains eight copies of each of these conserved regions. Southern analyses has demonstrated that sapA2 existed as a complete copy within the genome in all strains examined, although Northern (RNA) analysis has demonstrated that sapA2 was not expressed in the C. fetus strain from which it was cloned. Further Southern analyses revealed increasing sapA diversity as probes increasingly 3' within the ORF were used. Pulsed-field gel electrophoresis and then Southern blotting with the conserved N-terminal region of the sapA homologs as a probe showed that these genes were tightly clustered on the chromosome. Deletion mutagenesis revealed that the S-layer protein bound serospecifically to the C. fetus lipopolysaccharide via its conserved N-terminal region. These data indicated that the S-layer proteins shared functional activity in the conserved N terminus but diverged in a semiconservative manner for the remainder of the molecule. Variation in S-layer protein expression may involve rearrangement of complete gene copies from a single large locus containing multiple sapA homologs.
Dworkin
J
J
Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee 37232.
Tummuru
M K
MK
Blaser
M J
MJ
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D015139:Blotting, Southern; D002168:Campylobacter fetus / Q000737:chemistry*; D003001:Cloning, Molecular; D016521:Electrophoresis, Gel, Pulsed-Field; D008070:Lipopolysaccharides / Q000378:metabolism*; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data
D001426:Bacterial Proteins; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; C055764:S-layer proteins