Partial site-specific assignment of a uniformly C, N enriched membrane protein, light-harvesting complex 1 (LH1), by solid state NMR |
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Authors: | Lei Huang Ann E. McDermott |
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Affiliation: | a Key Laboratory of Molecular Engineering of Polymers of the Ministry of Education, Department of Macromolecular Science, Fudan University, Shanghai 200433, China b Columbia University, Department of Chemistry, 3000 Broadway, New York, NY 10027, USA |
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Abstract: | Partial site-specific assignments are reported for the solid state NMR spectra of light-harvesting complex 1, a 160 kDa integral membrane protein. The assignments were derived from 600 MHz 15N-13CO-13Cα and 15N-13Cα-13CX correlation spectra, using uniformly 13C, 15N enriched hydrated material, in an intact and precipitated form. Sequential assignments were verified using characteristic 15N-13Cα-13Cβ side chain chemical shifts observed in 3D experiments. Tertiary contacts found in 2D DARR spectra of the selectively 13C enriched sample provided further confirmatory evidence for the assignments. The assignments include the region of the Histidine ligands binding the Bacteriochlorophyll chromophore. The chemical shifts of Cα and Cβ resonances indicated the presence of typical α-helical secondary structure, consistent with previous studies. |
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Keywords: | RC, (reaction center) Bchl, (bacteriochlorophyll) LH1, (light-harvesting complex 1) LH2, (light-harvesting complex 2) LHC, (light-harvesting complex) HEPES, (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) βOG, (1-O-octyl-β- smallcaps" >d-glucopyranoside) MPD, (2-methyl-2,4-pentanediol) DSS, (2,2-Dimethyl-2-silapentane-5-sulfonate sodium salt) 1D, (one-dimensional) 2D, (two-dimensional) 3D, (three-dimensional) CP, (cross polarization) CW, (continuous waves) TPPM, (two pulses phase modulation) DARR, (dipolar assisted rotational resonance) RFDR, (radio frequency driven dipolar recoupling) DCP, (double cross polarization) DCP-RFDR, (double cross polarization transfer followed by radio frequency driven dipolar recoupling) DCP-DARR, (double cross polarization transfer followed by dipolar assisted rotational resonance polarization transfer) CαCO, (chemical shift correlation between carbon alpha and carbonyl) NCACX, (nitrogen to carbon alpha followed by carbon alpha to carbonyl and side chain carbon polarization transfer) NCOCA, (nitrogen to carbonyl followed by carbonyl to carbon alpha polarization transfer) TPPI, (time proportional phase incrementation) |
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