• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Magic angle spinning NMR of the protonated retinylidene Schiff base nitrogen in rhodopsin: Expression of ~(15)N-lysine- and ~(13)C-glycine-labeled opsin in a stable cell line
【24h】

Magic angle spinning NMR of the protonated retinylidene Schiff base nitrogen in rhodopsin: Expression of ~(15)N-lysine- and ~(13)C-glycine-labeled opsin in a stable cell line

机译：视紫红质中质子化视黄叉席夫碱氮的魔角旋转核磁共振：稳定细胞系中〜（15）N-赖氨酸和〜（13）C-甘氨酸标记的视蛋白的表达

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

The apoprotein corresponding to the mam- malian photoreceptor rhodopsin has been expressed by using suspension cultures of HEK293S cells in defined media that contained 6-~(15)N-lysine and 2-~(13)C-glycine. Typical yields were 1.5-1.8 mg/liter. Incorporation of 6-~(15)N-lysine was quantita- tive, whereas that of 2-~(13)C-glycine was about 60. The rho- dopsin pigment formed by binding of 11-cis retinal was spectrally indistinguishable from native bovine rhodopsin. Magic angle spinning (MAS) NMR spectra of labeled rho- dopsin were obtained after its incorporation into liposomes. The ~(15)N resonance corresponding to the protonated retinyli- dene Schiff base nitrogen was observed at 156.8 ppm in the MAS spectrum of 6-~(15)N-lysine-labeled rhodopsin. This chem- ical shift corresponds to an effective Schiff base-counterion distance of greater than 4 A, consistent with structural water in the binding site hydrogen bonded with the Schiff base nitrogen and the Glu-113 counterion. The present study demonstrates that structural studies of rhodopsin and other G protein-coupled receptors by using MAS NMR are feasible.
机译：通过使用HEK293S细胞在含有6-〜（15）N-赖氨酸和2-〜（13）C-甘氨酸的特定培养基中悬浮培养，已经表达了对应于哺乳动物感光体视紫红质的载脂蛋白。典型的产量为1.5-1.8mg /升。 6-〜（15）N-赖氨酸的掺入量是定量的，而2-〜（13）C-甘氨酸的掺入量约为60。通过结合11-顺式视网膜形成的视紫红质色素在光谱上与天然色素没有区别。牛视紫红质。标记的视紫红质掺入脂质体后，获得了幻角旋转（MAS）NMR光谱。在6-（（15）N-赖氨酸标记的视紫红质的MAS光谱中，在156.8ppm处观察到了对应于质子化视黄醛席夫碱氮的〜（15）N共振。这种化学位移相当于大于4 A的有效席夫碱抗衡距离，这与席夫碱氮和Glu-113抗衡离子结合的氢结合位点中的结构水一致。本研究表明，通过MAS NMR对视紫红质和其他G蛋白偶联受体进行结构研究是可行的。

著录项

相似文献

  • 外文文献
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号