Sequence analysis of the 47-kilodalton major integral membrane immunogen of Treponema pallidum.
The complete primary amino acid sequence for the 47-kilodalton (kDa) major integral membrane immunogen of Treponema pallidum subsp. pallidum was obtained by using a combined strategy of DNA sequencing (of the cloned gene in Escherichia coli) and N-terminal amino acid sequencing of the native (T. pallidum subsp. pallidum-derived) antigen. An open reading frame believed to encode the 47-kDa antigen comprised 367 amino acid codons, which gave rise to a calculated molecular weight for the corresponding antigen of 40,701. Of the 367 amino acids, 113 (31%) were sequenced by N-terminal amino acid sequencing of trypsin and hydroxylamine cleavage fragments of the native molecule isolated from T. pallidum subsp. pallidum; amino acid sequence data had a 100% correlation with that of the amino acid sequence predicted from DNA sequencing of the cloned gene in E. coli. Although no consensus sequences for the initiation of transcription or translation were readily identifiable immediately 5' to the putative methionine start codon, a 63-base-pair PstI fragment located 159 nucleotides upstream was required for expression of the 47-kDa antigen in E. coli. The 47-kDa antigen sequence did not reveal a typical leader sequence. The overall G+C content for the DNA corresponding to the structural gene was 53%. Hydrophilicity analysis identified at least one major hydrophilic domain of the protein near the N terminus of the molecule which potentially represents an immunodominant epitope. No repetitive primary sequence epitopes were found. The combined data provide the molecular basis for further structural and functional studies regarding the role of the antigen in the immunopathogenesis of treponemal disease.
Hsu
P L
PL
Department of Pathology, University of Texas Health Science Center, Houston 77030.
Chamberlain
N R
NR
Orth
K
K
Moomaw
C R
CR
Zhang
L Q
LQ
Slaughter
C A
CA
Radolf
J D
JD
Sell
S
S
Norgard
M V
MV
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000235:genetics / Q000302:isolation & purification*; D000954:Antigens, Surface / Q000235:genetics / Q000302:isolation & purification*; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification*; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000302:isolation & purification; D005798:Genes, Bacterial; D008565:Membrane Proteins / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000235:genetics / Q000302:isolation & purification; D014210:Treponema pallidum / Q000235:genetics / Q000276:immunology*
D000942:Antigens, Bacterial; D000954:Antigens, Surface; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008565:Membrane Proteins; D010446:Peptide Fragments; C058094:major integral membrane antigen, Treponema pallidum