KdpFABC reconstituted in Escherichia coli lipid vesicles: substrate dependence of the transport rate.
KdpFABC reconstituted in Escherichia coli lipid vesicles: substrate dependence of the transport rate.
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KdpFABC 在大肠杆菌脂囊泡中重建：转运速率的底物依赖性
DOI：
10.1021/bi5008244
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发表时间：
2014
期刊：
影响因子：
2.9
通讯作者：
Damnjanovic
中科院分区：
文献类型：
--
作者：
Damnjanovic
KdpFABC复合体在大肠埃希氏菌的囊泡中重组，并在生理条件下通过在与细胞质相对应的外部介质中加入ATP来激活离子泵。研究了不同底物对细胞转运速率的影响，并研究了ATP驱动的钾分泌。电压敏感染料DiSC3(5)检测到泵浦电流随膜电位的降低而降低。结果表明，高K+浓度对KdpFABC复合体有抑制作用。当K+浓度为140mM，最低浓度为∼时，泵浦电流降至最大值的25%。这一效应可以被细胞质一侧增加的镁离子浓度所抵消。这一现象可以用两个镁离子的Gouy-Chapman效应来解释，这些离子可能与0.8 mm的AK1/2结合，靠近进入结合部位的通道入口。这一因素保证了生理条件下K+释放的限速效应显着降低。此外，腺苷二磷酸和无机磷都能通过分别颠倒磷酸化步骤(Kiof151μM)和去磷酸化步骤(Kiof268μM)来降低泵的周转率。在DDM增溶KdpFABC络合物的情况下，转换条件下的活化能为55kJ/m ol，钒酸络合物的抑制常数为∼1μM，这与文献报道的其他P型ATPase的值一致。然而，在重组酶的情况下，观察到的显著差异必须归因于脂双层环境的影响。活化能增加2倍，而钒酸根对酶的抑制作用降低，只有66%的∼能被抑制，抑制常数增大到∼60μM。
KdpFABC complexes were reconstituted inEscherichia colilipid vesicles, and ion pumping was activated by addition of ATP to the external medium which corresponds to the cytoplasm under physiological conditions. ATP-driven potassium extrusion was studied in the presence of various substrates potentially influencing transport rate. The pump current was detected as a decrease of the membrane potential by the voltage-sensitive dye DiSC3(5). The results indicate that high cytoplasmic K+concentrations have an inhibitory effect on the KdpFABC complex. The pump current decreased to ∼25% of the maximal value at 140 mM K+and minimal Mg2+concentrations. This effect could be counteracted with increased Mg2+concentrations on the cytoplasmic side. This observation may be explained by the Gouy–Chapman effect of two Mg2+ions probably bound with aK1/2of 0.8 mM close to the entrance of the access channel to the binding sites. This factor ensures that under physiological conditions the rate-limiting effect of K+release is significantly reduced. Also both ADP and inorganic phosphate are able to reduce the turnover rate of the pump by reversing the phosphorylation step (Kiof 151 μM) and the dephosphorylation step (Kiof 268 μM), respectively. In the case of the DDM-solubilized KdpFABC complex, activation energy under turnover conditions was previously found to be 55 kJ/mol, and theo-vanadate inhibition constant is shown here to be ∼1 μM, which is in agreement with values reported for other P-type ATPases. In the case of the reconstituted enzyme, however, significant differences were observed that have to be assigned to effects of the lipid bilayer environment. The activation energy was increased by a factor of 2, whereas the inhibition byo-vanadate became reduced in a way that only ∼66% of the enzyme could be inhibited and the inhibition constant was increased to a value of ∼60 μM.
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大肠杆菌 Kdp-ATP 酶产生的电 K 转运。
影响因子：
2.9
作者：
K. Fendler;S. Dröse;K. Altendorf;E. Bamberg
通讯作者：
E. Bamberg
二价阳离子与钠和钾转运腺苷三磷酸酶的磷酸酶的结合。
影响因子：
4.8
作者：
Y. Fukushima;R. Post
通讯作者：
R. Post
钒酸盐与胃氢 (1 ) 转运、钾 (1 ) 刺激的 ATP 酶结合并抑制该酶的催化和转运活性
DOI：
--
发表时间：
1983
期刊：
影响因子：
--
作者：
L. Faller;E. Rabon;G. Sachs
通讯作者：
G. Sachs
SCH28080 是胃 H,K-ATP 酶的 K 竞争性抑制剂，结合在 M5-6 腔环附近，阻止 K 进入离子结合域。
影响因子：
2.9
作者：
Vagin,O;Denevich,S;Munson,K;Sachs,G
通讯作者：
Sachs,G
大肠杆菌中 ATP 驱动的钾转运系统结构蛋白的鉴定。
DOI：
--
发表时间：
1978
影响因子：
11.1
作者：
L. Laimins;David B. Rhoads;Karlheinz Altendorf;W. Epstein
通讯作者：
W. Epstein