Cofactor optimization: Difference between revisions
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Often an NMR sample can be improved by adding an appropriate cofactor. Here I have three examples: | Often an NMR sample can be improved by adding an appropriate cofactor such as a metal or a small organic molecule. | ||
Here I have three examples: | |||
#Addition of Zn to IscU | #Addition of Zn to IscU | ||
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In this example the effect of metal chelation on protein stability was probed by a 2D 1H–15N HSQC NMR experiment. A well dispersed spectra is indicative of global stability, collapsed spectra of instability<ref>Ramelota, T. et. al. (2004) Solution NMR structure of the iron-sulfur cluster assembly protetin U (IscU) with zinc bound the active site. Journal of Molecular Biology, 344, 567-583 </ref>. Due to the adverse effect of metal chelation on protein stability, and the high prevalence of zinc, it is likely that a subset of NESG targeted proteins with poorly resolved 2D 1H–15N HSQC spectra, may have inadvertently lost structural zinc during the protein sample preparation. | In this example the effect of metal chelation on protein stability was probed by a 2D 1H–15N HSQC NMR experiment. A well dispersed spectra is indicative of global stability, collapsed spectra of instability<ref>Ramelota, T. et. al. (2004) Solution NMR structure of the iron-sulfur cluster assembly protetin U (IscU) with zinc bound the active site. Journal of Molecular Biology, 344, 567-583 </ref>. Due to the adverse effect of metal chelation on protein stability, and the high prevalence of zinc, it is likely that a subset of NESG targeted proteins with poorly resolved 2D 1H–15N HSQC spectra, may have inadvertently lost structural zinc during the protein sample preparation. | ||
''Insert IscU NHSQC figures here'' | ''Insert IscU NHSQC figures here'' | ||
== References == | == References == | ||
<references /> | <references /> | ||
Revision as of 22:20, 18 November 2009
Often an NMR sample can be improved by adding an appropriate cofactor such as a metal or a small organic molecule.
Here I have three examples:
- Addition of Zn to IscU
- Addition of CoA to ?
- something else
Addition and removal of Zinc from Haemophilus influenae IscU
As many as 30% of proteins are predicted to be metal binding proteins and many of these may require metal for proper folding. In fact, a recent analysis of the PDB revealed that 14% of non-redundant entries contain stoichiometric amounts of six transition metals: Mn, Fe, Co, Ni, Cu or Zn [1]
Zincfinder by Passerini (2006)
In this example the effect of metal chelation on protein stability was probed by a 2D 1H–15N HSQC NMR experiment. A well dispersed spectra is indicative of global stability, collapsed spectra of instability[2]. Due to the adverse effect of metal chelation on protein stability, and the high prevalence of zinc, it is likely that a subset of NESG targeted proteins with poorly resolved 2D 1H–15N HSQC spectra, may have inadvertently lost structural zinc during the protein sample preparation.
Insert IscU NHSQC figures here
References
- ↑ Shi, W., Zhan, C., Ignatov, A., Manjasetty, B.A., Marinkovic, N., Sullivan, M., Huang, R., and Chance, M.R. (2005). Metalloproteomics: high-throughput structural and functional annotation of proteins in structural genomics. Structure 13, 1473-1486.
- ↑ Ramelota, T. et. al. (2004) Solution NMR structure of the iron-sulfur cluster assembly protetin U (IscU) with zinc bound the active site. Journal of Molecular Biology, 344, 567-583