Functional and antigenic properties of GlpO from Mycoplasma mycoides subsp. mycoides SC: characterization of a flavin adenine dinucleotide-binding site deletion mutant.
L-alpha-glycerophosphate oxidase (GlpO) plays a central role in virulence of Mycoplasma mycoides subsp. mycoides SC, a severe bacterial pathogen causing contagious bovine pleuropneumonia (CBPP). It is involved in production and translocation of toxic H(2)O(2) into the host cell, causing inflammation and cell death. The binding site on GlpO for the cofactor flavin adenine dinucleotide (FAD) has been identified as Gly(12)-Gly(13)-Gly(14)-Ile(15)-Ile(16)-Gly(17). Recombinant GlpO lacking these six amino acids (GlpODeltaFAD) was unable to bind FAD and was also devoid of glycerophosphate oxidase activity, in contrast to non-modified recombinant GlpO that binds FAD and is enzymatically active. Polyclonal monospecific antibodies directed against GlpODeltaFAD, similarly to anti-GlpO antibodies, neutralised H(2)O(2) production of M. mycoides subsp. mycoides SC grown in the presence of glycerol, as well as cytotoxicity towards embryonic calf nasal epithelial (ECaNEp) cells. The FAD-binding site of GlpO is therefore suggested as a valuable target site for the future construction of deletion mutants to yield attenuated live vaccines of M. mycoides subsp. mycoides SC necessary to efficiently combat CBPP.
Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Bern, Bern, Switzerland.
Daniela F
DF
Bischof
Edy M
EM
Vilei
Joachim
J
Frey
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial / Q000276:immunology* / Q000378:metabolism*; D001426:Bacterial Proteins / Q000276:immunology* / Q000378:metabolism; D001665:Binding Sites; D003001:Cloning, Molecular; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism*; D017353:Gene Deletion; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009176:Mycoplasma mycoides / Q000378:metabolism*; D011485:Protein Binding; D011817:Rabbits
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D005182:Flavin-Adenine Dinucleotide