Mijakovic Ivan, Poncet Sandrine, Boël Grégory, Mazé Alain, Gillet Sylvie, Jamet Emmanuel, Decottignies Paulette, Grangeasse Christophe, Doublet Patricia, Le Maréchal Pierre, Deutscher Josef
Laboratoire de Génétique des Microorganismes, CNRS/INRA/INA-PG UMR2585, 78850 Thiverval-Grignon, France.
EMBO J. 2003 Sep 15;22(18):4709-18. doi: 10.1093/emboj/cdg458.
Protein-tyrosine kinases regulating bacterial exopolysaccharide synthesis autophosphorylate on tyrosines located in a conserved C-terminal region. So far no other substrates have been identified for these kinases. Here we demonstrate that Bacillus subtilis YwqD not only autophosphorylates at Tyr-228, but that it also phosphorylates the two UDP-glucose dehydrogenases (UDP-glucose DHs) YwqF and TuaD at a tyrosine residue. However, phosphorylation of YwqF and TuaD occurs only in the presence of the transmembrane protein YwqC. The presumed intracellular C-terminal part of YwqC (last 50 amino acids) seems to interact with the tyrosine-kinase and to allow YwqD-catalysed phosphorylation of the two UDP-glucose DHs, which are key enzymes for the synthesis of acidic polysaccharides. However, only when phosphorylated by YwqD do the two enzymes exhibit detectable UDP-glucose DH activity. Dephosphorylation of P-Tyr-YwqF and P-Tyr-TuaD by the P-Tyr-protein phosphatase YwqE switched off their UDP-glucose DH activity. YwqE, which is encoded by the fourth gene of the B.subtilis ywqCDEF operon, also dephosphorylates P-Tyr-YwqD.
调节细菌胞外多糖合成的蛋白酪氨酸激酶在位于保守C端区域的酪氨酸上进行自身磷酸化。到目前为止，尚未鉴定出这些激酶的其他底物。在此我们证明，枯草芽孢杆菌YwqD不仅在Tyr-228处进行自身磷酸化，而且还在一个酪氨酸残基上磷酸化两个UDP-葡萄糖脱氢酶（UDP-葡萄糖DHs）YwqF和TuaD。然而，YwqF和TuaD的磷酸化仅在跨膜蛋白YwqC存在时发生。YwqC推测的细胞内C端部分（最后50个氨基酸）似乎与酪氨酸激酶相互作用，并允许YwqD催化两个UDP-葡萄糖DHs的磷酸化，这两个酶是酸性多糖合成的关键酶。然而，只有在被YwqD磷酸化后，这两种酶才表现出可检测到的UDP-葡萄糖DH活性。P-Tyr-蛋白磷酸酶YwqE对P-Tyr-YwqF和P-Tyr-TuaD的去磷酸化关闭了它们的UDP-葡萄糖DH活性。由枯草芽孢杆菌ywqCDEF操纵子的第四个基因编码的YwqE也使P-Tyr-YwqD去磷酸化。
