Protein structure and NMR spectroscopy Valine and Leucine
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A biochemist is studying the structure of a protein using NMR. He would like to measure the distance between hydrogens of the methyl groups of Val and Leu and other hydrogens in the molecule. Using a well established approach he is able to assign signals from the methyl groups of each Val and Leu residue in his protein. But then he recognized he did not know which methyl in each Val or Leu was pro-R and which was pro-S. He needed to know. He had two bottles of glucose. One contained uniformly 12C-enriched glucose and the other uniformly 13C-enriched glucose. He could express his protein in E. coli during growth on minimal media with glucose as the sole carbon source. He appreciated that he could tell if a methyl carbon labeled with 13C was adjacent (bonded to) another 13C or a 12C. The NMR spectrum of the 13C-12C case is one resonance line; the spectrum of the
13C-12C case is split into two resonance lines of half intensity and separated by 40 Hz (illustrated in the figure). Remember, 12C is invisible in the 13C NMR spectrum. Explain what is meant by pro-R and pro-S and why the methyl groups of Leu and Val are prochiral. Define the prochirality of the isopropyl groups of Val and Leu using the Cahn-Ingold-Prelog rules (the one's you learned in organic chemistry). How could the biochemist distinguish them using the NMR experiment above and the two bottles of glucose he has at his disposal? What general assumption would you have to make for this approach to work? How could you validate this assumption with the NMR data? Would these data be "absolute"? Why?
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Solution Summary
A problem that concerns isotopically labelled glucose and amino acid expression is discussed in detail.
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A biochemist is studying the structure of a protein using NMR. He
would like to measure the distance between hydrogens of the methyl groups of Val and Leu and other hydrogens in the molecule. Using a well
established approach he is able to assign signals from the
methyl groups of each Val and Leu residue in his protein. But then he recognized he did not know which methyl in each Val or Leu was pro-R and which was pro-S. He needed to know. He had two bottles of glucose. One contained uniformly 12C-
enriched glucose and the other uniformly 13C-enriched glucose. He could express his protein in E. coli during growth on minimal media with glucose as the sole carbon source. He appreciated that he could tell if a methyl carbon labeled with 13C was
adjacent (bonded to) another 13C or a 12C. The NMR spectrum of the 13C-12C case is one resonance line; the spectrum of the
13C-12C case is split into two resonance lines of ...
Purchase this Solution
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