Characterization of a bacterial gene encoding an autophosphorylating protein tyrosine kinase.
Acinetobacter johnsonii harbors a protein tyrosine kinase activity that is able to catalyze autophosphorylation, like a number of eukaryotic tyrosine kinases. A biochemical and genetic analysis of this enzyme was performed. Maximum phosphorylation in vitro was obtained by incubating the kinase for 2 min at pH 7.0 in the presence of 5 mM magnesium chloride. In contrast to eukaryotic enzymes, no inhibitory effect of genistein and no phosphorylation of synthetic substrates such as poly (Glu80 Tyr20) or angiotensin II were observed. The analysis of the bacterial kinase by two-dimensional gel electrophoresis revealed the presence of at least five isoforms, all phosphorylated exclusively at tyrosine, which supports the concept that autophosphorylation occurs at multiple sites within the protein. The cloning and nucleotide sequencing of the gene encoding this kinase were achieved, which represents the first molecular characterization of a gene of this type in bacteria. An open reading frame of 2199 nucleotides encoding a protein of 82,373 Da was detected. The analysis of the deduced amino acid sequence suggested a possible involvement of the enzyme in cell recognition and bacterial pathogenicity. In addition, the cloning and sequencing of the region immediately upstream of the gene encoding the kinase revealed a novel open reading frame of 426 nucleotides encoding a phosphotyrosine protein phosphatase of 16,217 Da, which indicates that autophosphorylation on tyrosine is a physiologically reversible reaction.
Grangeasse
C
C
Institut de Biologie et Chimie des Protéines, Centre National de la Recherche Scientifique, Lyon, France.
Doublet
P
P
Vaganay
E
E
Vincent
C
C
Deléage
G
G
Duclos
B
B
Cozzone
A J
AJ
D000150:Acinetobacter / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D005798:Genes, Bacterial; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000235:genetics* / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism
D004269:DNA, Bacterial; D011505:Protein-Tyrosine Kinases; D017027:Protein Tyrosine Phosphatases