Mattoo Abid R, Arora Amit, Maiti Souvik, Singh Yogendra
Institute of Genomics and Integrative Biology, Delhi, India.
FEBS J. 2008 Dec;275(24):6237-47. doi: 10.1111/j.1742-4658.2008.06748.x. Epub 2008 Nov 8.
Bacillus subtilis has three active tyrosine kinases, PtkA, PtkB and McsB, which play an important role in the physiology of the bacterium. Genome sequence analysis and biochemical experiments indicated that the ortholog of McsB, BAS0080, is the only active tyrosine kinase present in Bacillus anthracis. The autophosphorylation of McsB of B. anthracis was enhanced in the presence of an activator protein McsA (BAS0079), a property similar to that reported for B. subtilis. However, the process of enhanced phosphorylation of McsB in the presence of McsA remains elusive. To understand the activation mechanism of McsB, we carried out spectroscopic and calorimetric experiments with McsB and McsA. The spectroscopic results suggest that the binding affinity of Mg-ATP for McsB increased by one order from 10(3) to 10(4) in the presence of McsA. The calorimetric experiments revealed that the interaction between McsB and McsA is endothermic in nature, with unfavourable positive enthalpy (DeltaH) and favourable entropy (DeltaS) changes leading to an overall favourable free energy change (DeltaG). Kinetics of binding of both ATP and McsA with McsB showed low association rates (k(a)) and fast dissociation rates (k(d)). These results suggest that enhanced phosphorylation of McsB in the presence of McsA is due to increased affinity of ATP for McsB.
枯草芽孢杆菌有三种活性酪氨酸激酶，即PtkA、PtkB和McsB，它们在该细菌的生理过程中发挥着重要作用。基因组序列分析和生化实验表明，炭疽芽孢杆菌中McsB的直系同源物BAS0080是唯一存在的活性酪氨酸激酶。在激活蛋白McsA（BAS0079）存在的情况下，炭疽芽孢杆菌McsB的自磷酸化作用增强，这一特性与枯草芽孢杆菌报道的类似。然而，在McsA存在时McsB磷酸化增强的过程仍不清楚。为了了解McsB的激活机制，我们对McsB和McsA进行了光谱和量热实验。光谱结果表明，在McsA存在的情况下，Mg-ATP对McsB的结合亲和力从10³增加到10⁴，提高了一个数量级。量热实验表明，McsB和McsA之间的相互作用本质上是吸热的，具有不利的正焓变（ΔH）和有利的熵变（ΔS），导致总体上有利的自由能变化（ΔG）。ATP和McsA与McsB的结合动力学显示出低缔合速率（k(a)）和快速解离速率（k(d)）。这些结果表明，在McsA存在时McsB磷酸化增强是由于ATP对McsB的亲和力增加。
