Schlictman D, Kavanaugh-Black A, Shankar S, Chakrabarty A M
Department of Microbiology and Immunology, University of Illinois College of Medicine, Chicago 60612.
J Bacteriol. 1994 Oct;176(19):6023-9. doi: 10.1128/jb.176.19.6023-6029.1994.
Infection with mucoid, alginate-producing strains of Pseudomonas aeruginosa is the leading cause of mortality among patients with cystic fibrosis. Alginate production by P. aeruginosa is not constitutive but is triggered by stresses such as starvation. The algR2 (also termed algQ) gene has been previously identified as being necessary for mucoidy; an algR2 mutant strain is unable to produce alginate when grown at 37 degrees C. We show here that the levels of phosphorylated succinyl coenzyme A synthetase (Scs) and nucleoside diphosphate kinase (Ndk), which form a complex in P. aeruginosa, are reduced in the algR2 mutant. We were able to correlate the lower level of phosphorylated Scs with a decrease in Scs activity. Western blots (immunoblots) also showed a decreased level of Ndk in the algR2 mutant, but the presence of another kinase activity sensitive to Tween 20 provides the missing Ndk function. The effect of AlgR2 on tricarboxylic acid (TCA) cycle enzymes appears to be specific for Scs, since none of the other TCA cycle enzymes measured showed a significant decrease in activity. Furthermore, the ability of the algR2 mutant to grow on TCA cycle intermediates, but not glucose, is impaired. These data indicate that AlgR2 is responsible for maintaining proper operation of the TCA cycle and energy metabolism.
感染产黏液、藻酸盐的铜绿假单胞菌菌株是囊性纤维化患者死亡的主要原因。铜绿假单胞菌产生藻酸盐不是组成型的，而是由饥饿等应激因素触发的。藻R2（也称为藻Q）基因先前已被确定为形成黏液所必需的；藻R2突变株在37摄氏度生长时无法产生藻酸盐。我们在此表明，在铜绿假单胞菌中形成复合物的磷酸化琥珀酰辅酶A合成酶（Scs）和核苷二磷酸激酶（Ndk）的水平在藻R2突变体中降低。我们能够将较低水平的磷酸化Scs与Scs活性的降低相关联。蛋白质免疫印迹法（免疫印迹）也显示藻R2突变体中Ndk水平降低，但存在另一种对吐温20敏感的激酶活性提供了缺失的Ndk功能。藻R2对三羧酸（TCA）循环酶的作用似乎对Scs具有特异性，因为所检测的其他TCA循环酶均未显示活性有显著降低。此外，藻R2突变体在TCA循环中间产物而非葡萄糖上生长的能力受损。这些数据表明藻R2负责维持TCA循环和能量代谢的正常运作。
