Tiburzy H J, Zimmermann M, Oworah-Nkruma R, Berzborn R J
Lehrstuhl für Biochemie der Pflanzen, Fakultät für Biologie der Ruhr-Universität Bochum, Germany.
Z Naturforsch C J Biosci. 1999 Mar-Apr;54(3-4):230-8. doi: 10.1515/znc-1999-3-413.
Subunit II is one of the four nonidentical subunits of the membrane integral, proton-transporting moiety (CFo) of the chloroplast ATP synthase. In chloroplasts of spinach leaves, it is the only nuclear-encoded CFo subunit. It has been deduced that CFoII is not an additional subunit typical for photosynthetic organisms with no counterpart in E. coli, but equivalent to E. coli subunit b (Tiburzy, H.-J. and Berzborn, R. J. (1997), Z. Naturforsch. 52c, 789-798). Heterologous expression of subunit II was achieved by using the bacterial expression vector pT7-7. Recombinant subunit II (IIrec) does not integrate into the bacterial membrane nor does it precipitate into inclusion bodies. Gel filtration chromatography indicates that IIrec forms higher order aggregates. In three chromatographic steps approx. 10 mg of soluble IIrec of electrophoretic homogeneity are obtained from one liter of bacterial culture without using detergents. Thus, a eukaryotic membrane-anchored protein has been overexpressed in E. coli and has been purified in a soluble form.
亚基II是叶绿体ATP合酶膜整合质子转运部分（CFo）的四个不同亚基之一。在菠菜叶的叶绿体中，它是唯一由核编码的CFo亚基。据推测，CFoII并非光合生物特有的、在大肠杆菌中没有对应物的额外亚基，而是等同于大肠杆菌的亚基b（蒂布尔齐，H.-J.和贝茨伯恩，R. J.（1997年），《自然科学杂志》52c，789 - 798页）。通过使用细菌表达载体pT7 - 7实现了亚基II的异源表达。重组亚基II（IIrec）既不整合到细菌膜中，也不沉淀形成包涵体。凝胶过滤色谱表明IIrec形成了高阶聚集体。在三个色谱步骤中，无需使用去污剂，从一升细菌培养物中可获得约10毫克电泳纯的可溶性IIrec。因此，一种真核膜锚定蛋白已在大肠杆菌中过表达，并以可溶形式纯化出来。
