Identification of a bacterial inhibitor of protein kinases. Mechanism and role in host cell invasion.
We show that Escherichia coli produce a factor that inhibits the activity of tyrosine and serine/threonine protein kinases. The factor is a protein found in the periplasmic compartment and is also secreted into the culture medium. Using a particle concentration fluorescence immunoassay specific for tyrosine kinase activity and inhibition of the tyrosine kinase p56(lck), we purified this factor to apparent homogeneity. Analysis of trypsin-digested fragments by mass spectrometry identified the inhibitor as the bacterial periplasmic protein UDP-sugar hydrolase, an enzyme with potent and nonspecific 5'-nucleotidase activity. Overexpression of the enzyme in bacteria leads to coordinate increases in both 5'-nucleotidase and p56(lck) inhibitory activity, confirming the identity of the inhibitor. The kinase inhibitory activity appears to be due to the formation of adenosine, which we show is inhibitory for p56(lck), cAMP-dependent protein kinase, and casein kinase. Overexpression of UDP-sugar hydrolase leads to an increase in the recovery of enteropathogenic E. coli following infection of HeLa cell monolayers and corresponding alterations in tyrosine-phosphorylated host proteins. These results suggest that UDP-sugar hydrolase may be an important factor affecting host cell function following intracellular bacterial infection.
Berger
S A
SA
Wellesley Hospital Research Institute, Toronto, Ontario, Canada M4Y 1J3.
Rowan
K
K
Morrison
H D
HD
Ziltener
H J
HJ
D000241:Adenosine / Q000494:pharmacology*; D000818:Animals; D047388:Casein Kinases; D002478:Cells, Cultured / Q000382:microbiology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D004926:Escherichia coli / Q000737:chemistry* / Q000472:pathogenicity; D006801:Humans; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D051379:Mice; D010727:Phosphoric Diester Hydrolases / Q000235:genetics / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins / Q000378:metabolism; D019061:src-Family Kinases / Q000037:antagonists & inhibitors
D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D019061:src-Family Kinases; D047388:Casein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D010727:Phosphoric Diester Hydrolases; D000241:Adenosine