Two-dimensional electrophoresis for analysis of Mycobacterium tuberculosis culture filtrate and purification and characterization of six novel proteins.
Culture filtrate from Mycobacterium tuberculosis contains molecules which promote high levels of protective immunity in animal models of subunit vaccination against tuberculosis. We have used two-dimensional electrophoresis for analysis and purification of six novel M. tuberculosis culture filtrate proteins (CFPs): CFP17, CFP20, CFP21, CFP22, CFP25, and CFP28. The proteins were tested for recognition by M. tuberculosis-reactive memory cells from different strains of inbred mice and for their capacity to induce a skin test response in M. tuberculosis-infected guinea pigs. CFP17, CFP20, CFP21 and CFP25 induced both a high gamma interferon release and a strong delayed-type hypersensitivity response, and CFP21 was broadly recognized by different strains of inbred mice. N-terminal sequences were obtained for the six proteins, and the corresponding genes were identified in the Sanger M. tuberculosis genome database. In parallel we established a two-dimensional electrophoresis reference map of short-term culture filtrate components and mapped novel proteins as well as already-known CFP.
Department of TB Immunology, Statens Serum Institut, Copenhagen, Denmark.
K
K
Weldingh
I
I
Rosenkrands
S
S
Jacobsen
P B
PB
Rasmussen
M J
MJ
Elhay
P
P
Andersen
D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D002478:Cells, Cultured; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D005260:Female; D015870:Gene Expression; D005798:Genes, Bacterial; D006168:Guinea Pigs; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D017421:Sequence Analysis
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C113716:CFP17 protein, Mycobacterium tuberculosis; C113717:CFP20 protein, Mycobacterium tuberculosis; C113719:CFP21 protein, Mycobacterium tuberculosis; C113721:CFP22 protein, Mycobacterium tuberculosis; C113750:CFP25 protein, Mycobacterium tuberculosis