Demonstration of two protein kinases in extracts of Legionella micdadei.
Protein kinases I (PK I) and II (PK II) were purified 253- and 13.5-fold, respectively, from an extract of sonically disrupted cells of Legionella micdadei by ion-exchange chromatography on QAE-Sephadex, by histone affinity chromatography, and by HPLC-gel filtration chromatography. Both enzymes catalysed the phosphorylation of calf thymus histones, with a Km of 2.7 mg ml-1 for PK I and 2.9 mg ml-1 for PK II. Histone H2b was the best protein kinase substrate for both PK I and PK II. The pH optima were 6.8 and 7.0 for PK I and PK II respectively. The Km for ATP was 0.29 mM for PK I and 0.33 mM for PK II. PK II activity was stimulated by either cAMP or cGMP, whereas PK I was inhibited by both cyclic nucleotides. The activity of PK I was unaffected by addition of calmodulin, diacylglycerol and mixtures of Ca2+ and acidic phospholipids, but these additions increased PK II activity threefold. The activity of PK II was stimulated by spermine and spermidine, but PK I was inhibited by these compounds. PK I and PK II were both strongly inhibited by heparin.
Saha
A K
AK
Department of Microbiology, Biochemistry and Molecular Biology, University of Pittsburgh School of Medicine, PA 15261.
Dowling
J N
JN
Mukhopadhyay
N K
NK
Glew
R H
RH
D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D000242:Cyclic AMP / Q000494:pharmacology; D006152:Cyclic GMP / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007875:Legionella / Q000201:enzymology*; D011494:Protein Kinases / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity
D000242:Cyclic AMP; D011494:Protein Kinases; D006152:Cyclic GMP