Zhang Li, Jiang Wangshu, Nan Jie, Almqvist Jonas, Huang Yafei
Department of Cell and Molecular Biology, Uppsala University, Uppsala Biomedical Center, Box 596, 75124 Uppsala, Sweden.
Department of Cell and Molecular Biology, Uppsala University, Uppsala Biomedical Center, Box 596, 75124 Uppsala, Sweden.
Biochim Biophys Acta. 2014 Jul;1838(7):1809-16. doi: 10.1016/j.bbamem.2014.03.003. Epub 2014 Mar 19.
The Escherichia coli inner membrane protein CysZ mediates the sulfate uptake subsequently utilized for the synthesis of sulfur-containing compounds in cells. Here we report the purification and functional characterization of CysZ. Using Isothermal Titration Calorimetry, we have observed interactions between CysZ and its putative substrate sulfate. Additional sulfur-containing compounds from the cysteine synthesis pathway have also been analyzed for their abilities to interact with CysZ. Our results suggest that CysZ is dedicated to a specific pathway that assimilates sulfate for the synthesis of cysteine. Sulfate uptake via CysZ into E. coli whole cells and proteoliposome offers direct evidence of CysZ being able to mediate sulfate uptake. In addition, the cysteine synthesis pathway intermediate sulfite can interact directly with CysZ with higher affinity than sulfate. The sulfate transport activity is inhibited in the presence of sulfite, suggesting the existence of a feedback inhibition mechanism in which sulfite regulates sulfate uptake by CysZ. Sulfate uptake assays performed at different extracellular pH and in the presence of a proton uncoupler indicate that this uptake is driven by the proton gradient.
大肠杆菌内膜蛋白CysZ介导硫酸盐的摄取，随后这些硫酸盐被用于细胞中含硫化合物的合成。在此，我们报告了CysZ的纯化及功能特性。使用等温滴定量热法，我们观察到CysZ与其假定底物硫酸盐之间的相互作用。还分析了来自半胱氨酸合成途径的其他含硫化合物与CysZ相互作用的能力。我们的结果表明，CysZ专门参与一条将硫酸盐同化为半胱氨酸合成的特定途径。通过CysZ将硫酸盐摄取到大肠杆菌全细胞和蛋白脂质体中，为CysZ能够介导硫酸盐摄取提供了直接证据。此外，半胱氨酸合成途径的中间体亚硫酸盐可以比硫酸盐以更高的亲和力直接与CysZ相互作用。在亚硫酸盐存在的情况下，硫酸盐转运活性受到抑制，这表明存在一种反馈抑制机制，其中亚硫酸盐调节CysZ对硫酸盐的摄取。在不同细胞外pH值下以及在质子解偶联剂存在的情况下进行的硫酸盐摄取试验表明，这种摄取是由质子梯度驱动的。
