Published on Web 07/17/2004
Amino Acid-Type Edited NMR Experiments for Methyl-Methyl Distance Measurement in 13C-Labeled Proteins He´ le` ne Van Melckebeke, Jean-Pierre Simorre, and Bernhard Brutscher* Contribution from the Institut de Biologie Structurale, Jean-Pierre Ebel CNRS-CEA-UJF, 41, rue Jules Horowitz, 38027 Grenoble Cedex, France Received February 24, 2004; E-mail:
[email protected] Abstract: New NMR experiments are presented for the measurement of methyl-methyl distances in 13Clabeled proteins from a series of amino acid-type separated 2D or 3D NOESY spectra. Hadamard amino acid-type encoding of the proximal methyl groups provides the high spectral resolution required for unambiguous methyl-methyl NOE assignment, which is particularly important for fast global fold determination of proteins. The experiments can be applied to a wide range of protein systems, as exemplified for two small proteins, ubiquitin and MerAa, and the 30 kDa BRP-Blm complex.
Introduction
Liquid-state NMR spectroscopy has been established as a powerful method for structure determination of proteins. The development of many multidimensional triple-resonance NMR experiments over the past decade, combined with advanced isotope-labeling strategies, has significantly enlarged the scope of NMR problems that can now be studied by NMR. On one hand, TROSY-type experiments1 have increased the size of molecules amenable to an NMR investigation. Recently, nearcomplete backbone assignments have been reported for the 723residue malate synthase G protein from Escherichia coli,2 the 110 kDa homo-octameric protein 7,8-dihydroneolpterin aldolase,3 and the membrane proteins OmpX and OmpA.4 On the other hand, the development of new acquisition schemes has sped up NMR data collection.5 Such techniques are especially promising in the context of fast NMR fold determination of moderately sized proteins (
