CaMKII uses GTP as a phosphate donor for both substrate and autophosphorylation.

CaMKII uses GTP as a phosphate donor for both substrate and autophosphorylation.
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DOI:
10.1016/j.bbrc.2009.10.107
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发表时间:
2009-12-25
影响因子:
3.1
通讯作者:
Griffith, Leslie C.
Griffith, Leslie C.
中科院分区:
生物学4区
文献类型:
--
作者:
Bostrom, S. Lynn;Dore, Justin;Griffith, Leslie C.

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绝大多数丝氨酸/苏氨酸蛋白激酶对ATP的偏好超过GTP作为磷酸供体。CK 2(酪蛋白激酶2)是这一规则的例外,在这项研究中,我们调查是否钙/钙调蛋白依赖性蛋白激酶II(CaMKII)具有相同的扩展核苷酸范围。使用果蝇酶,我们已经表明,CaMKII使用Mg 2 +GTP,与Mg 2 +ATP相比,具有更高的Km和Vmax。Mn ~(2+)取代Mg ~(2+)导致GTP的Km值降低,同时几乎消除了CaMK Ⅱ利用ATP的能力。用大鼠αCaMKII获得了这些类似的结果,表明使用GTP的能力是CaMKII的一般性质。发现Mg 2 +ATP和Mg 2 +GTP之间的Vmax差异是由于ADP是磷酸化的有效抑制剂,而GDP具有适度的影响。无论是通过部分蛋白水解还是质谱分析,ATP和GTP自磷酸化位点之间均无差异。蝇头提取物的磷酸化显示,无论使用Mg 2 +ATP还是Mg 2 +GTP,类似的蛋白质都是CaMKII的底物。这一新的信息证实了CaMKII可以使用ATP和GTP,并为研究这种激酶的调节开辟了新的途径。
The vast majority of serine/threonine protein kinases have a strong preference for ATP over GTP as a phosphate donor. CK2 (Casein kinase 2) is an exception to this rule and in this study we investigate whether calcium/calmodulin-dependent protein kinase II (CaMKII) has the same extended nucleotide range. Using the Drosophila enzyme, we have shown that CaMKII uses Mg2+GTP with a higher Km and Vmax compared to Mg2+ATP. Substitution of Mn2+ for Mg2+ resulted in a much lower Km for GTP, while nearly abolishing the ability of CaMKII to use ATP. These similar results were obtained with rat αCaMKII, showing the ability to use GTP to be a general property of CaMKII. The Vmax difference between Mg2+ATP and Mg2+GTP was found to be due to the fact that ADP is a potent inhibitor of phosphorylation, while GDP has modest effects. There were no differences found between sites autophosphorylated by ATP and GTP, either by partial proteolysis or mass spectrometry. Phosphorylation of fly head extract revealed that similar proteins are substrates for CaMKII whether using Mg2+ATP or Mg2+GTP. This new information confirms that CaMKII can use both ATP and GTP, and opens new avenues for the study of regulation of this kinase.
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