Solid state NMR spectral properties and partial sequential assignment of a membrane protein, the light harvesting complex 1 (LH1)

Recent progress in solid state NMR indicates that it is a promising technique for high resolution protein structure determination. One of its advantages is its applicability to systems that are difficult to study by solution NMR or X-ray crystallography, e.g. membrane proteins. In this work, we applied solid state NMR methods to the LH1 complex of purple bacteria Rhodobacter sphaeroides, which forms a ring-like structure involving 16 pairs of alphabeta peptides, with each alphabeta consisting of 106 amino acids. Three-dimensional NMR experiments were performed at high magnetic field to establish inter- and intra-residue correlations. Partial sequential assignments were derived from those experiments.; Factors that affect solid state NMR spectral qualities were investigated. Different detergents (betaOG, DMPC and DHPC) and precipitants (PEG400, PEG6000 and MPD) were tested to identify optimal protein sample conditions. Temperature was critical in obtaining high-resolution spectra, as well as in maintaining protein stability. The optimal temperature balances those two demands. Uniformly 13C, 15N isotopic enrichment facilitated the site-specific backbone assignment, while the use of selective 13C enrichment greatly reduced spectral congestion, helping to identify additional resonance assignments that might be ambiguous in the spectra of uniformly isotopical enriched samples. The spectra of selectively isotopical enriched proteins also provided important structural information based on assignment of long-range contacts. Cofactors (bacteriochlorophyll and carotenoid) play fundamental structural and functional roles in the photosynthetic system. NMR signals for cofactors were observed in two-dimensional 13C- 13C and 13C-15N spectra, which assisted in establishing protein-pigment interactions in light harvesting proteins.