abstract

• A particular class of tyrosine-kinases sharing no structural similarity with eukaryotic tyrosine-kinases has been evidenced in a large array of bacterial species. These bacterial tyrosine-kinases are able to autophosphorylate on a C-terminal tyrosine-rich motif. Their autophosphorylation has been shown to play a crucial role in the biosynthesis or export of capsular polysaccharide. The analysis of the first crystal structure of the staphylococcal tyrosine kinase CapB2 associated with the activating domain of the transmembrane modulator CapA1 had brought conclusive explanation for both the autophosphorylation and activation processes. In order to explain why CapA1 activates CapB2 more efficiently than its cognate transmembrane modulator CapA2, we solved the crystal structure of CapA2B2 and compared it with the previously published structure of CapA1B2. This structural analysis did not provide the expected clues about the activation discrepancy observed between the two modulators. Staphylococcus aureus also encodes for a CapB2 homologue named CapB1 displaying more than 70%25 sequence similarity and being surprisingly nearly unable to autophosphorylate. We solved the crystal structure of CapA1B1 and carefully compare it with the structure of CapA1B2. The active sites of both proteins are highly conserved and the biochemical characterization of mutant proteins engineered to test the importance of small structural discrepancies identified between the two structures did not explain the inactivity of CapB1. We thus tested if CapB1 could phosphorylate other protein substrates or hydrolyze ATP. However, no activity could be detected in our in vitro assays. Taken together, these data question about the biological role of the homologous protein pairs CapA1/CapB1 and CapA2/CapB2 and we discuss about several possible interpretations. © 2013 Gruszczyk et al.

authors

• Aumont-Nicaise M.
• Bechet E.
• Fleurie A.
• Freton C.
• Grangeasse C.
• Gruszczyk J.
• Gueguen-Chaignon V.
• Morera S.
• Nessler S.
• Nourikyan J.
• Olivares Illana Vanesa

publication date

• 2013-01-01

published in

• PLoS ONE  Journal

keywords

• asparagine; bacterial enzyme; protein tyrosine kinase; protein tyrosine kinase CapA1; protein tyrosine kinase CapA2; protein tyrosine kinase CapB1; protein tyrosine kinase CapB2; unclassified drug; amino acid sequence; amino terminal sequence; article; autophosphorylation; carboxy terminal sequence; comparative study; controlled study; crystal structure; crystallization; differential scanning calorimetry; enzyme activation; enzyme activity; enzyme phosphorylation; Escherichia coli; expression vector; molecular cloning; nonhuman; protein function; protein hydrolysis; protein motif; protein purification; site directed mutagenesis; Staphylococcus aureus; Amino Acid Sequence; Bacterial Proteins; Catalytic Domain; Crystallography, X-Ray; Escherichia coli; Gene Expression Regulation, Bacterial; Isoenzymes; Models, Molecular; Molecular Sequence Data; Phosphorylation; Protein Binding; Protein Structure, Tertiary; Protein-Tyrosine Kinases; Recombinant Proteins; Sequence Alignment; Staphylococcus aureus; Structural Homology, Protein

Digital Object Identifier (DOI)

• 10.1371/journal.pone.0075958

PubMed ID

• 24146800

start page

end page

volume

• 8

issue

• 10