Full engagement of liganded maltose-binding protein stabilizes a semi-open ATP-binding cassette dimer in the maltose transporter.
Full engagement of liganded maltose-binding protein stabilizes a semi-open ATP-binding cassette dimer in the maltose transporter.
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DOI：
10.1111/mmi.13165
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发表时间：
2015-12
影响因子：
3.6
通讯作者：
Davidson AL
中科院分区：
文献类型：
--
作者：
Alvarez FJ;Orelle C;Huang Y;Bajaj R;Everly RM;Klug CS;Davidson AL
MalFGK2是一种三磷酸腺苷结合盒(ABC)转运蛋白，介导麦芽糖/麦芽糊精进入大肠杆菌。周质麦芽糖结合蛋白(MBP)将麦芽糖输送到跨膜亚基(MalFG)，并刺激细胞质核苷酸结合亚基(Malk二聚体)的ATPase活性。这种MBP刺激的ATPase活性不依赖于麦芽糖对洗涤剂胶束中纯化转运蛋白的作用。然而，当转运蛋白在膜双层中重组时，只有连接形式的MBP才能有效地刺激其活性。为了探讨麦芽糖的刺激机制，用电子顺磁共振(EPR)研究了转运蛋白与MBP在纳米盘和洗涤剂中的相互作用。我们发现，核苷酸促进了麦芽糖结合的MBP的两个叶与MalFGK2的完全结合，并稳定了半开放的Malk二聚体。当转运蛋白在膜上时，与麦芽糖结合的MBP促进麦芽糖向半开放状态的转变，而这种调节不需要洗涤剂中的麦芽糖。我们认为，麦芽糖结合的MBP对半开放的MalK2构象的稳定是麦芽糖转运与体内ATP水解偶联的关键，因为它促进了Malk二聚体从开放构象到半开放构象的发展，从而可以进一步水解ATP。
MalFGK2 is an ATP-binding cassette (ABC) transporter that mediates the uptake of maltose/maltodextrins into Escherichia coli. A periplasmic maltose-binding protein (MBP) delivers maltose to the transmembrane subunits (MalFG) and stimulates the ATPase activity of the cytoplasmic nucleotide-binding subunits (MalK dimer). This MBP-stimulated ATPase activity is independent of maltose for purified transporter in detergent micelles. However, when the transporter is reconstituted in membrane bilayers, only the liganded form of MBP efficiently stimulates its activity. To investigate the mechanism of maltose stimulation, electron paramagnetic resonance (EPR) spectroscopy was used to study the interactions between the transporter and MBP in nanodiscs and in detergent. We found that full engagement of both lobes of maltose-bound MBP unto MalFGK2 is facilitated by nucleotides and stabilizes a semi-open MalK dimer. Maltose-bound MBP promotes the transition to the semi-open state of MalK when the transporter is in the membrane, whereas such regulation does not require maltose in detergent. We suggest that stabilization of the semi-open MalK2 conformation by maltose-bound MBP is key to the coupling of maltose transport to ATP hydrolysis in vivo, because it facilitates the progression of the MalK dimer from the open to the semi-open conformation, from which it can proceed to hydrolyze ATP.