Mode of Interaction of the Signal-Transducing Protein EIIA(Glc) with the Maltose ABC Transporter in the Process of Inducer Exclusion.

Mode of Interaction of the Signal-Transducing Protein EIIA(Glc) with the Maltose ABC Transporter in the Process of Inducer Exclusion.
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诱导子排除过程中信号转导蛋白EIIA(Glc)与麦芽糖ABC转运蛋白的相互作用模式

DOI:
10.1021/acs.biochem.6b00721
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发表时间:
2016
期刊:
影响因子:
2.9
通讯作者:
Schneider
Schneider
中科院分区:
生物学3区
文献类型:
--
作者:
Wuttge;Timachi;Bordignon;Schneider

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大肠杆菌和沙门氏菌中用于摄取葡萄糖的磷酸烯醇式丙酮酸磷酸转移酶系统的酶IIAGlc(EIIAGlc)通过与核苷酸结合和水解亚基MalK相互作用来抑制麦芽糖ATP结合盒转运蛋白(MalE-FGK 2),这一过程称为诱导物排除。我们已经研究了EIIAGlc与MalK二聚体在蛋白脂质体中通过半胱氨酸交联的结合。结果证明,EIIAGl的结合位点I与MalK的N-末端亚结构域接触,而结合位点II相对接近C-末端(调节)亚结构域,与晶体结构一致[Chen,S.,奥尔德姆,M. L.,戴维森,A. L.,和Chen,J.(2013)Nature 499,364 - 368]。此外,发现EIIAGlc结合MalK二聚体,而不管其构象状态如何。缺失抑制所需的EIIAGlc的两亲性N-末端肽,减少了交联产物的形成。使用自旋标记的转运蛋白变体和EPR光谱,我们证明了EIIAGl通过抑制MalK二聚体的ATP依赖性闭合来进行转运循环。
Enzyme IIAGlc(EIIAGlc) of the phosphoenolpyruvate phosphotransferase system for the uptake of glucose inEscherichia coliandSalmonellainhibits the maltose ATP-binding cassette transporter (MalE-FGK2) by interaction with the nucleotide-binding and -hydrolyzing subunit MalK, a process termed inducer exclusion. We have investigated binding of EIIAGlcto the MalK dimer by cysteine cross-linking in proteoliposomes. The results prove that the binding site I of EIIAGlcis contacting the N-terminal subdomain of MalK while the binding site II is relatively close to the C-terminal (regulatory) subdomain, in agreement with a crystal structure [Chen, S., Oldham, M. L., Davidson, A. L., and Chen, J. (2013) Nature 499, 364−368]. Moreover, EIIAGlcwas found to bind to the MalK dimer regardless of its conformational state. Deletion of the amphipathic N-terminal peptide of EIIAGlc, which is required for inhibition, reduced formation of cross-linked products. Using a spin-labeled transporter variant and EPR spectroscopy, we demonstrate that EIIAGlcarrests the transport cycle by inhibiting the ATP-dependent closure of the MalK dimer.
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通过针对 MalK 亚基的单克隆抗体对鼠伤寒沙门氏菌的麦芽糖 ATP 结合盒转运蛋白进行功能表征。
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