Functional incorporation of chimeric b subunits into F1Fo ATP synthase.
Functional incorporation of chimeric b subunits into F1Fo ATP synthase.
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嵌合 b 亚基功能性并入 F1Fo ATP 合酶。
DOI：
10.1128/jb.00191-07
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发表时间：
2007
影响因子：
3.2
通讯作者：
Cain,BrianD
中科院分区：
文献类型：
--
作者：
Claggett,ShaneB;Grabar,TammyBohannon;Dunn,StanleyD;Cain,BrianD
F1FoATP合酶通过旋转机制起作用。酶的外围茎作为定子，保持f1扇区及其催化位点不受转子运动的影响。在大肠杆菌中，外周茎是相同蛋白亚基的同二聚体，但光合细菌有两个不同的亚基的开放阅读框，被认为形成异二聚体/b '外周茎。通过对长胞热聚球菌带基因序列的替换，构建了嵌合亚基基因。大肠杆菌uncf基因，编码该亚基。重组基因分别在体外和体外表达。碰撞碰撞应变KM2 （Δb）。虽然不是所有的嵌合亚基都被纳入F1FoATP合成酶复合物，但表达chimericbE39-I86orb ' e39 - i86的质粒都能够在功能上与菌株KM2互补（Δb）。表达这些亚单位的菌株比表达e39 - d53orbl54 - i86亚单位的嵌合片段较小的细胞生长得更好，表明基因内抑制。一般来说，嵌合亚基以t为模型。在体外实验中，长藤b亚基比b亚基更稳定。菌株KM2 （Δb）中be39 - i86和b ' e39 - i86亚基的共表达产生了含有由这两个亚基组成的异二聚体外周茎的f1fo复合物。
F1FoATP synthases function by a rotary mechanism. The enzyme's peripheral stalk serves as the stator that holds the F1sector and its catalytic sites against the movement of the rotor. InEscherichia coli, the peripheral stalk is a homodimer of identicalbsubunits, but photosynthetic bacteria have open reading frames for two differentb-like subunits thought to form heterodimericb/b′ peripheral stalks. Chimericbsubunit genes have been constructed by substituting sequence from theThermosynechococcus elongatus bandb′ genes in theE. coli uncFgene, encoding thebsubunit. The recombinant genes were expressed alone and in combination in theE. colideletion strain KM2 (Δb). Although not all of the chimeric subunits were incorporated into F1FoATP synthase complexes, plasmids expressing either chimericbE39-I86orb′E39-I86were capable of functionally complementing strain KM2 (Δb). Strains expressing these subunits grew better than cells with smaller chimeric segments, such as those expressing theb′E39-D53orbL54-I86subunit, indicating intragenic suppression. In general, the chimeric subunits modeled on theT. elongatus bsubunit proved to be more stable than theb′ subunit in vitro. Coexpression of thebE39-I86andb′E39-I86subunits in strain KM2 (Δb) yielded F1Focomplexes containing heterodimeric peripheral stalks composed of both subunits.