Molecular characterization of the major outer membrane protein of Haemophilus somnus.
The major outer membrane protein (MOMP) of Haemophilus somnus shows antigenic and molecular mass diversity that forms the basis of a preliminary grouping system for H. somnus strains. In this study, the gene encoding MOMP of H. somnus strain 8025 was cloned in three overlapping fragments by PCR techniques, and then sequenced. The gene consists of a 1164-bp open reading frame encoding a deduced 380-amino acid protein with a 19-amino acid signal sequence, giving a mature protein with a calculated molecular mass of 39,913 Da. Significant homology was found between MOMP and porin protein sequences of bacteria in Pasteurellaceae species. When expressed in Escherichia coli, the protein from the MOMP gene directed by the T7 promoter was identical in size (approximately 40 kDa) to native MOMP and reacted with MOMP-specific antibodies. Comparisons of the MOMP gene sequences from six unrelated strains of H. somnus to that of strain 8025 revealed that the genes of three MOMP type 1 strains were highly conserved with that of strain 8025 in length and sequence. However, two MOMP type 3c strains and one MOMP type 3a strain differed markedly from the MOMP of strain 8025 in their 3'-terminal halves. Their deduced MOMP amino acid sequences differed in sequence (3c, 80.5 and 82.7% identity; 3a, 62.4% identity) and in length (3c, 384 and 376; 3a, 316), indicating that the molecular differences are the basis of antigenicity and molecular mass differences of H. somnus MOMP. In the predicted MOMP secondary structure, the variable sequences primarily mapped to putative surface-exposed loops, and a variable and surface-exposed epitope of MOMP-specific antibody was identified in the seventh-largest loop. These findings are useful for understanding the structural and immunological characteristics of H. somnus.
Khan
Md Shahidur R
MS
National Institute of Animal Health, 3-1-5 Kannondai, Tsukuba, Ibaraki 305-0856, Japan.
Tanaka
Akinori
A
Ide
Hisahiro
H
Hoshinoo
Kaori
K
Hanafusa
Yasuko
Y
Tagawa
Yuichi
Y
D000595:Amino Acid Sequence; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D001483:Base Sequence; D002417:Cattle; D002418:Cattle Diseases / Q000382:microbiology*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D018604:Epitope Mapping; D004926:Escherichia coli / Q000235:genetics; D006192:Haemophilus Infections / Q000662:veterinary; D044138:Haemophilus somnus / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016133:Polymerase Chain Reaction / Q000662:veterinary; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial