Abstract:

Nuclear magnetic resonance (NMR) has emerged as a powerful tool for determining three-dimensional structures of proteins in solution. The major part of this dissertation describes the efforts to deal with the core steps involved in NMR-base protein structure determination: NMR data collection, NMR data analysis, structure calculation and refinement.

NMR data collection has been recognized as one of the major bottlenecks of NMR-base structure determination due to the necessarily long time. Chapter 2 and 3 describe two NMR data collection and analysis protocols for high-quality protein structure determination based on Reduced Dimensionality (RD) and G-matrix Fourier transform (GFT) NMR, respectively. The rapidly collected RD & GFT NMR data enabled high-quality structure determination of four structural genomics target proteins in high-throughput. Chapter 5 introduces the program UBNMR which was developed to facilitate NMR data analysis in general and RD & GFT NMR data pre-processing and data analysis in particular. RD and GFT NMR based protocols, with the aid of UBNMR, are expected to greatly impact on the NMR-based structural biology and structural genomics.

In addition to rapid NMR data collection and analysis, structure calculation and refinement are also pivotal for obtaining the high-quality protein structures. Chapter 5 describes the analysis of the newly implemented simultaneous GFT NOESY to obtain an accurate and precise initial structural fold. Chapter 6 presents a protein structure refinement strategy using NOE data collected in supercooled water at low temperatures.