Retinylidene Ligand Structure in Bovine Rhodopsin, Metarhodopsin-I, and 10-Methylrhodopsin from Internuclear Distance Measurements Using 13C-Labeling and 1-D Rotational Resonance MAS NMR† Export

@article{citeulike:1811289, abstract = {PMID: 10471281 Rhodopsin is the G-protein coupled photoreceptor that initiates the rod phototransduction cascade in the vertebrate retina. Using specific isotope enrichment and magic angle spinning (MAS) NMR, we examine the spatial structure of the C10C11C12C13C20 motif in the native retinylidene chromophore, its 10-methyl analogue, and the predischarge photoproduct metarhodopsin-I. For the rhodopsin study 11-Z-[10,20-13C2]- and 11-Z-[11,20-13C2]-retinal were synthesized and incorporated into bovine opsin while maintaining a natural lipid environment. The ligand is covalently bound to Lys296 in the photoreceptor. The C10C20 and C11C20 distances were measured using a novel 1-D CP/MAS NMR rotational resonance experimental procedure that was specifically developed for the purpose of these measurements [Verdegem, P. J. E., Helmle, M., Lugtenburg, J., and de Groot, H. J. M. (1997) J. Am. Chem. Soc. 119, 169]. We obtain r10,20 = 0.304 +- 0.015 nm and r11,20 = 0.293 +- 0.015 nm, which confirms that the retinylidene is 11-Z and shows that the C10C13 unit is conformationally twisted. The corresponding torsional angle is about 44o as indicated by CarParrinello modeling studies. To increase the nonplanarity in the chromophore, 11-Z-[10,20-13C2]-10-methylretinal and 11-Z-[(10-CH3),13-13C2]-10-methylretinal were prepared and incorporated in opsin. For the resulting analogue pigment r10,20 = 0.347 +- 0.015 nm and r(10-CH3),13 = 0.314 +- 0.015 nm were obtained, consistent with a more distorted chromophore. The analogue data are in agreement with the induced fit principle for the interaction of opsin with modified retinal chromophores. Finally, we determined the intraligand distances r10,20 and r11,20 also for the photoproduct metarhodopsin-I, which has a relaxed all-E structure. The results (r10,20 0.435 nm and r11,20 = 0.283 +- 0.015 nm) fully agree with such a relaxed all-E structure, which further validates the 1-D rotational resonance technique for measuring intraligand distances and probing ligand structure. As far as we are aware, these results represent the first highly precise distance determinations in a ligand at the active site of a membrane protein. Overall, the MAS NMR data indicate a tight binding pocket, well defined to bind specifically only one enantiomer out of four possibilities and providing a steric complement to the chromophore in an ultrafast (200 fs) isomerization process.}, address = {Leiden Institute of Chemistry, Gorlaeus Laboratories, The Netherlands.}, author = {Verdegem, P. J. E. and Bovee-Geurts, P. H. M. and de Grip, W. J. and Lugtenburg, J. and de Groot, H. J. M.}, citeulike-article-id = {1811289}, citeulike-linkout-0 = {http://dx.doi.org/10.1021/bi983014e}, citeulike-linkout-1 = {http://pubs.acs.org/doi/abs/10.1021/bi983014e}, citeulike-linkout-2 = {http://view.ncbi.nlm.nih.gov/pubmed/10471281}, citeulike-linkout-3 = {http://www.hubmed.org/display.cgi?uids=10471281}, day = {1}, doi = {10.1021/bi983014e}, issn = {0006-2960}, journal = {Biochemistry}, keywords = {activation, induced\_fit, metai, nmr, retinal, rhodopsin}, month = {August}, number = {35}, pages = {11316--11324}, posted-at = {2009-11-06 10:42:33}, priority = {2}, title = {Retinylidene Ligand Structure in Bovine Rhodopsin, Metarhodopsin-I, and 10-Methylrhodopsin from Internuclear Distance Measurements Using 13C-Labeling and 1-D Rotational Resonance MAS NMR{\dag}}, url = {http://dx.doi.org/10.1021/bi983014e}, volume = {38}, year = {1999} }

TY - JOUR ID - citeulike:1811289 L3 - citeulike-article-id:1811289 N2 - PMID: 10471281 Rhodopsin is the G-protein coupled photoreceptor that initiates the rod phototransduction cascade in the vertebrate retina. Using specific isotope enrichment and magic angle spinning (MAS) NMR, we examine the spatial structure of the C10C11C12C13C20 motif in the native retinylidene chromophore, its 10-methyl analogue, and the predischarge photoproduct metarhodopsin-I. For the rhodopsin study 11-Z-[10,20-13C2]- and 11-Z-[11,20-13C2]-retinal were synthesized and incorporated into bovine opsin while maintaining a natural lipid environment. The ligand is covalently bound to Lys296 in the photoreceptor. The C10C20 and C11C20 distances were measured using a novel 1-D CP/MAS NMR rotational resonance experimental procedure that was specifically developed for the purpose of these measurements [Verdegem, P. J. E., Helmle, M., Lugtenburg, J., and de Groot, H. J. M. (1997) J. Am. Chem. Soc. 119, 169]. We obtain r10,20 = 0.304 +- 0.015 nm and r11,20 = 0.293 +- 0.015 nm, which confirms that the retinylidene is 11-Z and shows that the C10C13 unit is conformationally twisted. The corresponding torsional angle is about 44o as indicated by CarParrinello modeling studies. To increase the nonplanarity in the chromophore, 11-Z-[10,20-13C2]-10-methylretinal and 11-Z-[(10-CH3),13-13C2]-10-methylretinal were prepared and incorporated in opsin. For the resulting analogue pigment r10,20 = 0.347 +- 0.015 nm and r(10-CH3),13 = 0.314 +- 0.015 nm were obtained, consistent with a more distorted chromophore. The analogue data are in agreement with the induced fit principle for the interaction of opsin with modified retinal chromophores. Finally, we determined the intraligand distances r10,20 and r11,20 also for the photoproduct metarhodopsin-I, which has a relaxed all-E structure. The results (r10,20 0.435 nm and r11,20 = 0.283 +- 0.015 nm) fully agree with such a relaxed all-E structure, which further validates the 1-D rotational resonance technique for measuring intraligand distances and probing ligand structure. As far as we are aware, these results represent the first highly precise distance determinations in a ligand at the active site of a membrane protein. Overall, the MAS NMR data indicate a tight binding pocket, well defined to bind specifically only one enantiomer out of four possibilities and providing a steric complement to the chromophore in an ultrafast (200 fs) isomerization process. IS - 35 JF - Biochemistry SN - 0006-2960 AD - Leiden Institute of Chemistry, Gorlaeus Laboratories, The Netherlands. EP - 11324 TI - Retinylidene Ligand Structure in Bovine Rhodopsin, Metarhodopsin-I, and 10-Methylrhodopsin from Internuclear Distance Measurements Using 13C-Labeling and 1-D Rotational Resonance MAS NMR† VL - 38 SP - 11316 KW - activation KW - induced_fit KW - metai KW - nmr KW - retinal KW - rhodopsin AU - Verdegem, PJE AU - Bovee-Geurts, PHM AU - de Grip, WJ AU - Lugtenburg, J AU - de Groot, HJM PY - 1999/08/01/ UR - http://dx.doi.org/10.1021/bi983014e ER -