研究目的
To develop and apply modified heteronuclear correlation experiments using solid-state photo-CIDNP for assigning NMR resonances of protons bound to photochemically active cofactors in photosynthetic reaction centers.
研究成果
The photo-CIDNP MAS-J-HMQC experiment is effective for mapping proton chemical shifts in photosynthetic cofactors, providing selective spectra without protein backbone interference. It aids in understanding electronic structures and potential intermolecular polarization transfers, with good agreement between experimental and theoretical shifts. Future work should focus on higher MAS frequencies and uniform labeling for complete assignments.
研究不足
The experiments suffer from low sensitivity due to fast transverse relaxation times and homonuclear dipolar couplings; transfer efficiency depends on s delays and relaxation times, with potential for improvement at higher MAS frequencies. Some signals remain unassigned, possibly due to spin diffusion or accessory cofactors.
1:Experimental Design and Method Selection:
Utilized photo-CIDNP MAS-J-HMQC and MAS-J-HSQC pulse sequences to transfer light-induced hyperpolarization from 13C to 1H nuclei via J-couplings, with homonuclear decoupling (PMLG5-S2) during transfer delays and t1 evolution.
2:Sample Selection and Data Sources:
Selectively 13C-labeled photosynthetic reaction centers from Rhodobacter sphaeroides WT, grown with 3-, 4-, or 5-13C-d-aminolevulinic acid, isolated and quinone-blocked.
3:List of Experimental Equipment and Materials:
Bruker AVANCE III spectrometers at
4:4 T and 20 T, 4-mm MAS probes, sapphire and zirconia rotors, xenon-arc lamp for illumination, temperature control units, and software (Bruker TopSpin 2, MNova 12). Experimental Procedures and Operational Workflow:
Samples frozen in dark, spun at specific MAS frequencies (e.g., 7518 Hz), illuminated continuously, NMR experiments performed with optimized pulse sequences, parameters (e.g., s delays, PMLG decoupling), and data acquisition with multiple scans and increments.
5:Data Analysis Methods:
Processed with Fourier transformation, window functions (sine bell), zero-filling, and referenced to solid L-tyrosine hydrochloride; compared with DFT calculations and solution NMR data.
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