Previous Article | Next Article ![]()
Journal of Bacteriology, November 1999, p. 6615-6622, Vol. 181, No. 21
Department of Microbiology and Immunology,
University of Illinois College of Medicine, Chicago, Illinois 60612
Received 19 May 1999/Accepted 18 August 1999
Pseudomonas aeruginosa is an opportunistic pathogen
that causes infections in eye, urinary tract, burn, and
immunocompromised patients. We have cloned and characterized a
serine/threonine (Ser/Thr) kinase and its cognate phosphoprotein
phosphatase. By using oligonucleotides from the conserved regions of
Ser/Thr kinases of mycobacteria, an 800-bp probe was used to screen
P. aeruginosa PAO1 genomic library. A 20-kb cosmid clone
was isolated, from which a 4.5-kb DNA with two open reading frames
(ORFs) were subcloned. ORF1 was shown to encode Ser/Thr phosphatase
(Stp1), which belongs to the PP2C family of phosphatases. Overlapping
with the stp1 ORF, an ORF encoding Hank's type Ser/Thr
kinase was identified. Both ORFs were cloned in pGEX-4T1 and
expressed in Escherichia coli. The overexpressed proteins
were purified by glutathione-Sepharose 4B affinity chromatography and
were biochemically characterized. The Stk1 kinase is 39 kDa and
undergoes autophosphorylation and can phosphorylate eukaryotic histone
H1. A site-directed Stk1 (K86A) mutant was shown to be incapable of
autophosphorylation. A two-dimensional phosphoamino acid analysis of
Stk1 revealed strong phosphorylation at a threonine residue and weak
phosphorylation at a serine residue. The Stp1 phosphatase is 27 kDa and
is an Mn2+-, but not a Ca2+- or a
Mg2+-, dependent Ser/Thr phosphatase. Its activity is
inhibited by EDTA and NaF, but not by okadaic acid, and is similar to
that of PP2C phosphatase.
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Characterization of a Hank's Type Serine/Threonine
Kinase and Serine/Threonine Phosphoprotein Phosphatase in
Pseudomonas aeruginosa
*
Corresponding author. Mailing address: Department of
Microbiology and Immunology (M/C 790), University of Illinois at
Chicago College of Medicine, 835 S. Wolcott Ave., Chicago, IL 60612. Phone: (312) 996-4586. Fax: (312) 996-6415. E-mail:
Ananda.Chakrabarty{at}uic.edu.
This article has been cited by other articles:
| Appl. Environ. Microbiol. | Infect. Immun. | Eukaryot. Cell |
|---|---|---|
| Mol. Cell. Biol. | J. Virol. | Microbiol. Mol. Biol. Rev. |
| ALL ASM JOURNALS |