Measuring translational diffusion coefficients of peptides and proteins by PFG-NMR using band-selective RF pulses.
Molecular translational self-diffusion, a measure of diffusive motion, provides information on the effective molecular hydrodynamic radius, as well as information on the properties of media or solution through which the molecule diffuses. Protein translational diffusion measured by pulsed-field gradient nuclear magnetic resonance (PFG-NMR) has seen increased application in structure and interaction studies, as structural changes or protein-protein interactions are often accompanied by alteration of their effective hydrodynamic radii. Unlike the analysis of complex mixtures by PFG-NMR, for monitoring changes of protein translational diffusion under various conditions, such as different stages of folding/unfolding, a partial region of the spectrum or even a single resonance is sufficient. We report translational diffusion coefficients measured by PFG-NMR with a modified stimulated echo (STE) sequence where band-selective pulses are employed for all three (1)H RF pulses. Compared with conventional non-selective sequence, e.g. the BPP-LED sequence, the advantage of this modified band-selective excitation short transient (BEST) version of STE (BEST-STE) sequence is multi-fold, namely: (1) potential sensitivity gain as in generalized BEST-based sequences, (2) water suppression is no longer required as the magnetization of solvent water is not perturbed during the measurement, and (3) dynamic range problems due to the presence of intense resonances from molecules other than the protein or peptide of interest, such as non-deuterated detergent micelles, are avoided.
Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Melbourne, VIC, 3010, Australia, shyao@unimelb.edu.au.
Shenggen
S
Yao
Daniel K
DK
Weber
Frances
F
Separovic
David W
DW
Keizer
D016229:Amyloid beta-Peptides / Q000737:chemistry* / Q000378:metabolism*; D004058:Diffusion; D009682:Magnetic Resonance Spectroscopy; D008823:Micelles; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism*; D012997:Solvents / Q000494:pharmacology; D013501:Surface-Active Agents / Q000494:pharmacology; D013696:Temperature; D051028:bcl-2-Associated X Protein / Q000737:chemistry* / Q000378:metabolism*
D016229:Amyloid beta-Peptides; D008823:Micelles; D010446:Peptide Fragments; D012997:Solvents; D013501:Surface-Active Agents; C075222:amyloid beta-protein (1-42); D051028:bcl-2-Associated X Protein