Genetic manipulation of immunoglobulin binding proteins of Haemophilus somnus.
The relationship of the 76kDa immunoglobulin binding, surface antigen (p76) of Haemophilus somnus to the high molecular weight immunoglobulin binding proteins (HMW IgBPs) was investigated. The kanamycin resistance gene from pLS88 was used via homologous recombination with allelic exchange to replace a portion of the gene encoding IgBPs of H. somnus strain 8025. Recombinants were shown by Western immunoblotting to express and secrete truncated antigens of approximately 200kDa and not to produce p76. The truncated HMW IgBP variants retained the ability to bind bovine IgG2 by the Fc portion as demonstrated by Western immunoblotting against IgG2 anti-DNP. This data indicated that the deleted 1.8kb BglII fragment was not required for secretion or immunoglobulin Fc binding by the HMW IgBPs but was required for expression of the downstream p76 gene. Functional studies showed that, in addition to Fc binding of IgG2 to truncated HMW IgBPs, the mutant strain 8025 Kan1 was equally resistant to killing by mouse complement but less virulent than the wild type parent (8025) in a mouse septicemia model of H. somnus infection. However, mutant strain 8025 Kan1 did adhere less well than the wild type to bovine pulmonary artery endothelial cells. It is probable that p76 and the missing peptides of the HMW IgBPs play a role in this aspect of virulence and perhaps other aspects.
Sanders
Jerry D
JD
Department of Pathology, University of California, San Diego Medical Center, 200 West Arbor Drive, San Diego, CA 92103-8416, USA.
Bastida-Corcuera
Felix D
FD
Arnold
Karen F
KF
Wunderlich
Annette C
AC
Corbeil
Lynette B
LB
D000818:Animals; D016470:Bacteremia / Q000382:microbiology / Q000401:mortality; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D004730:Endothelium, Vascular; D017353:Gene Deletion; D006190:Haemophilus / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D006192:Haemophilus Infections / Q000382:microbiology / Q000401:mortality; D007074:Immunoglobulin G / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D023421:Models, Animal; D011485:Protein Binding; D011651:Pulmonary Artery; D011995:Recombination, Genetic
D001426:Bacterial Proteins; D007074:Immunoglobulin G; D008565:Membrane Proteins; C114756:TM9SF2 protein, human