Loss of activity in the secreted form of Escherichia coli haemolysin caused by an rfaP lesion in core lipopolysaccharide assembly.
A transposon mutant of Escherichia coli 5K was isolated which reduced 10- to 50-fold the secreted extracellular haemolytic activity of cells carrying the complete hlyCABD operon while leaving unaffected the intracellular haemolytic activity and the levels of intracellular and extracellular haemolysin protein, HlyA. The transposon insertion was identified within the rfaP gene (required for attachment of phosphate-containing substituents to the lipopolysaccharide inner core), and extracellular haemolytic activity was restored in trans by the intact rfaP gene. The loss in cytolytic activity of the secreted HlyA protein was not related to the HlyC-directed acylation of the protoxin. Activity of the secreted toxin was restored by chaotropic agents and during rate-zonal centrifugation the mutant-secreted HlyA migrated as a larger species than the wild type. The results indicate that the rfaP mutation affects the aggregation behaviour of the active toxin during or following the signal peptide-independent secretion process.
Stanley
P L
PL
Cambridge University Department of Pathology, UK.
Diaz
P
P
Bailey
M J
MJ
Gygi
D
D
Juarez
A
A
Hughes
C
C
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D006460:Hemolysin Proteins / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000378:metabolism*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional
D001426:Bacterial Proteins; D017931:DNA Primers; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D006460:Hemolysin Proteins; C036507:Hlya protein, E coli; D008070:Lipopolysaccharides