Flagellar proteins and type III-exported virulence factors are the predominant proteins secreted into the culture media of Salmonella typhimurium

We analysed all major proteins secreted into culture media from Salmonella typhimurium. Proteins in culture supernatants were collected by trichloroac etic acid precipitation, separated in SDS-polyacrylamide gels and analysed by amino acid sequencing. Wild-type strain SJW1103 cells typically gave ris e to nine bands in SDS gels: 89, 67, 58, 52, 50, 42, 40, 35 and (sometimes) 28 kDa. A search of the sequences in the available databases revealed that they were either flagellar proteins or virulence factors. Six of them were flagella specific: FlgK or HAP1 (58 kDa), FliC or flagellin (52 kDa), FliD or HAP2 (50 kDa), FlgE or hook protein (42 kDa), FlgL or HAP3 (35 kDa) and FlgD or hook-cap protein (28 kDa). The other four bands were specific for virulence factors: SipA (89 kDa), SipB (67 kDa), SipC (42 kDa) and InvJ (40 kDa). The 42 kDa band was a mixture of FlgE and SipC. We also analysed sec reted proteins from more than 30 flagellar mutants, and they were categoriz ed into four groups according to their band patterns: wild type, mot type, polyhook type and master gene type. Virulence factors were constantly secre ted at a higher level in all flagellar mutants except a Delta mot (motAB de letion) mutant, in which the amounts were greatly reduced. A new morphologi cal pathway of flagellar biogenesis including protein secretion is presente d.