Use of residual dipolar couplings in structural analysis of protein-ligand complexes by solution NMR spectroscopy.
Investigation of structure-function relationships in protein complexes, specifically protein-ligand interactions, carry great significance in elucidating the structural and mechanistic bases of molecular recognition events and their role in regulating cell processes. Nuclear magnetic resonance (NMR) spectroscopy is one of the leading structural and analytical techniques in in-depth studies of protein-ligand interactions. Recent advances in NMR methodology such as transverse relaxation-optimized spectroscopy (TROSY) and residual dipolar couplings (RDCs) measured in liquid crystalline alignment medium, offer a viable alternative to traditional nuclear Overhauser enhancement (NOE)-based approaches for structure determination of large protein complexes. RDCs provide a way to constrain the relative orientation of two molecules in complex with each other by aligning their independently determined order tensors. The potential for utilization of RDCs can be extended to proteins with multiple ligands or even multimeric protein-ligand complexes, where symmetry properties of the protein can be taken advantage of. Availability of effective RDC data collection and analysis protocols can certainly aid this process by their incorporation into structure calculation protocols using intramolecular and intermolecular orientational restraints. This chapter discusses in detail some of these protocols including methods for sample preparation in liquid crystalline media, NMR experiments for RDC data collection, as well as software tools for RDC data analysis and protein-ligand complex structure determination.
Jain
Nitin U
NU
Cellular and Molecular Biology Department, University of Tennessee, 37996-0840, Knoxville, TN, USA. njain@utk.edu
D055664:Electrochemical Techniques; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D037601:Mannose-Binding Lectin / Q000737:chemistry; D046912:Multiprotein Complexes / Q000737:chemistry; D017351:N-Acetylglucosaminyltransferases / Q000737:chemistry; D009844:Oligosaccharides / Q000737:chemistry; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry*; D012996:Solutions; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000737:chemistry
D008024:Ligands; D037601:Mannose-Binding Lectin; D046912:Multiprotein Complexes; D009844:Oligosaccharides; D011506:Proteins; D012996:Solutions; C028569:oligomannoside; D014537:Uridine Diphosphate N-Acetylglucosamine; D017351:N-Acetylglucosaminyltransferases; C052254:alpha-1,6-mannosylglycoprotein beta 1,6-N-acetylglucosaminyltransferase