微线圈 NMR 监测整合膜蛋白 OmpW 在去污剂胶束中复性条件的优化。
Micro-coil NMR to monitor optimization of the reconstitution conditions for the integral membrane protein OmpW in detergent micelles.
机构信息
Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
出版信息
J Biomol NMR. 2012 Oct;54(2):129-33. doi: 10.1007/s10858-012-9658-x. Epub 2012 Aug 14.
Abstract
Optimization of aqueous solutions of the integral membrane protein (IMP) OmpW for NMR structure determination has been monitored with micro-coil NMR, which enables the acquisition of NMR spectra using only micrograms of protein and detergent. The detergent 30-Fos (2-undecylphosphocholine) was found to yield the best 2D [(15)N, (1)H]-TROSY correlation NMR spectra of [(2)H, (15)N]-labeled OmpW. For the OmpW structure determination we then optimized the 30-Fos concentration, the sample temperature and long-time stability, and the deuteration level of the protein. Some emerging guidelines for reconstitution of β-barrel integral membrane proteins in structural biology are discussed.
摘要
采用微线圈 NMR 监测了整体膜蛋白（IMP）OmpW 的水溶液的优化，该方法仅使用微克级的蛋白质和去污剂即可获得 NMR 谱。发现去污剂 30-Fos（2-十一烷基磷酸胆碱）可产生最佳的二维 [(15)N，(1)H]-TROSY 相关 NMR 谱 [(2)H，(15)N]-标记的 OmpW。然后，我们针对 OmpW 结构测定优化了 30-Fos 浓度、样品温度和长时间稳定性以及蛋白质的氘化水平。讨论了一些用于结构生物学中β桶整体膜蛋白重建的新准则。
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