Optimization of NMR spectroscopy of encapsulated proteins dissolved in low viscosity fluids

Nathaniel V. Nucci, Bryan S. Marques, Sabrina Bédard, Jakob Dogan, John M. Gledhill, Veronica R. Moorman, Ronald W. Peterson, Kathleen G. Valentine, Alison L. Wand, A. Joshua Wand

Research output: Contribution to journalArticlepeer-review

15 Scopus citations

Abstract

Comprehensive application of solution NMR spectroscopy to studies of macromolecules remains fundamentally limited by the molecular rotational correlation time. For proteins, molecules larger than 30 kDa require complex experimental methods, such as TROSY in conjunction with isotopic labeling schemes that are often expensive and generally reduce the potential information available. We have developed the reverse micelle encapsulation strategy as an alternative approach. Encapsulation of proteins within the protective nano-scale water pool of a reverse micelle dissolved in ultra-low viscosity nonpolar solvents overcomes the slow tumbling problem presented by large proteins. Here, we characterize the contributions from the various components of the protein-containing reverse micelle system to the rotational correlation time of the encapsulated protein. Importantly, we demonstrate that the protein encapsulated in the reverse micelle maintains a hydration shell comparable in size to that seen in bulk solution. Using moderate pressures, encapsulation in ultra-low viscosity propane or ethane can be used to magnify this advantage. We show that encapsulation in liquid ethane can be used to reduce the tumbling time of the 43 kDa maltose binding protein from ~23 to ~10 ns. These conditions enable, for example, acquisition of TOCSY-type data resolved on the adjacent amide NH for the 43 kDa encapsulated maltose binding protein dissolved in liquid ethane, which is typically impossible for proteins of such size without use of extensive deuteration or the TROSY effect.

Original languageEnglish (US)
Pages (from-to)421-430
Number of pages10
JournalJournal of Biomolecular NMR
Volume50
Issue number4
DOIs
StatePublished - Aug 2011

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Spectroscopy

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