SITE DIRECTED ISOTOPE LABELING OF MEMBRANE PROTEINS

膜蛋白的定点同位素标记

基本信息

项目摘要

The SIR provided L-[4- 13 ClAspartic Acid; 2g L-[1,5_13C 2]Glutamic Acid; 0.5g FTIR difference spectroscopy can be used to probe conformational changes at the level of individual amino acid residues in a membrane protein. However, a key problem limiting progress is the absence of a general method for the incorporation of isotope labels at specific positions in a protein. Until recently this was not possible except through chemical synthesis, which is generally limited to polypeptides of 50 residues. We have recently introduced a new method which circumvents this limitation. Termed site-directed isotope labeling (SDIL), it is based on the use of suppressor tRNAs and in vitro synthesis. In the case of bacteriorhodopsin, the light- driven proton pump, we have used this method to label several tyrosine and peptide carbonyl positions. Combined with FrIR-difference spectroscopy, this has allowed us to pinpoint structural activity in Tyr residue 185 which may serve as a hinge for conformational changes in that protein. Our goal is now to extend these methods so that they can be used to label routinely any residue in bacteriorhodopsin; can be applied to other proteins and can be used in conjunction with biophysical techniques such as solid state NMR, neutron diffraction and FTIR. For this purpose new methods will be developed for both enzymatic and chemical arninoacylation of suppressor tRNAs to optimize in vitro expression, isolation and refolding and to characterize SDIL proteins spectroscopically. An extensive set of SDIL bacteriorhodopsin analogs will be produced including labels placed in Asp, Glu, Pro, Thr, Ser and Trp positions. These studies will directly lead to a detailed picture of how this protein functions. Similar research will be conducted on sensory rhodopsin I, the phototactic receptor in Halobacterium. salinarium.
SIR提供L-[4- 13] ClAspartic Acid;2 g

项目成果

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KENNETH J ROTHSCHILD其他文献

KENNETH J ROTHSCHILD的其他文献

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{{ truncateString('KENNETH J ROTHSCHILD', 18)}}的其他基金

Melanopsin Signal Transduction Studied by FTIR Spectroscopy
通过 FTIR 光谱研究黑视蛋白信号转导
  • 批准号:
    8132900
  • 财政年份:
    2010
  • 资助金额:
    $ 2.13万
  • 项目类别:
Melanopsin Signal Transduction Studied by FTIR Spectroscopy
通过 FTIR 光谱研究黑视蛋白信号转导
  • 批准号:
    7987558
  • 财政年份:
    2010
  • 资助金额:
    $ 2.13万
  • 项目类别:
Melanopsin Signal Transduction Studied by FTIR Spectroscopy
通过 FTIR 光谱研究黑视蛋白信号转导
  • 批准号:
    8271423
  • 财政年份:
    2010
  • 资助金额:
    $ 2.13万
  • 项目类别:
FTIR STUDY OF SIGNAL TRANSDUCTION IN SENSORY RHODOPSINS
感觉视紫红质信号转导的 FTIR 研究
  • 批准号:
    6873519
  • 财政年份:
    2005
  • 资助金额:
    $ 2.13万
  • 项目类别:
FTIR STUDY OF SIGNAL TRANSDUCTION IN SENSORY RHODOPSINS
感觉视紫红质信号转导的 FTIR 研究
  • 批准号:
    7175444
  • 财政年份:
    2005
  • 资助金额:
    $ 2.13万
  • 项目类别:
FTIR STUDY OF SIGNAL TRANSDUCTION IN SENSORY RHODOPSINS
感觉视紫红质信号转导的 FTIR 研究
  • 批准号:
    7342112
  • 财政年份:
    2005
  • 资助金额:
    $ 2.13万
  • 项目类别:
FTIR STUDY OF SIGNAL TRANSDUCTION IN SENSORY RHODOPSINS
感觉视紫红质信号转导的 FTIR 研究
  • 批准号:
    7007312
  • 财政年份:
    2005
  • 资助金额:
    $ 2.13万
  • 项目类别:
FTIR Study of Signal Transduction in Sensory Rhodopsins
感觉视紫红质信号转导的 FTIR 研究
  • 批准号:
    7737309
  • 财政年份:
    2005
  • 资助金额:
    $ 2.13万
  • 项目类别:
SITE DIRECTED ISOTOPE LABELING OF MEMBRANE PROTEINS
膜蛋白的定点同位素标记
  • 批准号:
    6309035
  • 财政年份:
    2000
  • 资助金额:
    $ 2.13万
  • 项目类别:
SITE DIRECTED ISOTOPE LABELING OF MEMBRANE PROTEINS
膜蛋白的定点同位素标记
  • 批准号:
    6251973
  • 财政年份:
    1997
  • 资助金额:
    $ 2.13万
  • 项目类别:

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