Enomoto H, Unemoto T, Nishibuchi M, Padan E, Nakamura T
Laboratory of Membrane Biochemistry, Faculty of Pharmaceutical Sciences, Chiba University, 1-33 Yayoi-cho, Inage-ku, Chiba 263, Japan.
Biochim Biophys Acta. 1998 Mar 6;1370(1):77-86. doi: 10.1016/s0005-2736(97)00245-9.
NhaB, an Na+/H+ antiporter, of Vibrio alginolyticus is a 528-amino-acid protein. Hydropathy profile-based computer analysis predicted that the NhaB might contain up to 13 membrane-spanning domains. To examine this hypothesis, we applied the phoA fusion method to the cloned nhaB gene. Eighteen plasmid-borne nhaB-phoA fusion genes were constructed in Escherichia coli cells and the alkaline phosphatase activity and expression level of the fusion proteins analyzed. These results and the results obtained with additional constructs indicated that V. alginolyticus NhaB has a unique topology consisting of nine transmembrane segments with the N-terminus in the cytoplasm and the C-terminus in the periplasm.
溶藻弧菌的 NhaB 是一种 Na⁺/H⁺逆向转运蛋白，由 528 个氨基酸组成。基于亲水性图谱的计算机分析预测，NhaB 可能含有多达 13 个跨膜结构域。为了验证这一假设，我们将 phoA 融合方法应用于克隆的 nhaB 基因。在大肠杆菌细胞中构建了 18 个质粒携带的 nhaB-phoA 融合基因，并分析了融合蛋白的碱性磷酸酶活性和表达水平。这些结果以及通过其他构建体获得的结果表明，溶藻弧菌 NhaB 具有独特的拓扑结构，由九个跨膜片段组成，N 端位于细胞质中，C 端位于周质中。
