Nuclear magnetic resonance studies of rhodopsin analogues derived from fluorinated retinals

Abstract

The compounds 9-cis- and all-trans-10-fluororetinal have been resynthesized with higher yields. Additionally, by direct irradiation of the all-trans-10-fluororetinal in acetonitrile, four other geometric isomers of 10-fluororetinal (including the sterically hindered 7-cis and 11-cis isomers) were obtained. The characterization of the geometry of these isomers and all synthetic intermediates was primarily based on 1H-NMR spectroscopy. For compounds in the 10-fluororetinal and 12-fluororetinal series, the observed 19F-NMR chemical shift trends were explained in terms of the conformation about the C8,C9 and C10,C11 single bonds respectively. The compounds 9-cis- and all-trans-β-ionylidene-fluoroacetaldehyde were synthetic intermediates to 10-fluororetinals. 7-Cis isomers of these C15 triene aldehydes were prepared by selective photosensitized isomerization. The ring orientation in three C15 triene aldehyde isomers (7-cis, 9-cis, and 7,9-dicis), relative to that of the dienal chain, was determined by means of magnetic resonance measurements of the nuclear Overhauser effect (NOE) and long-range coupling constants. The experimental results yield a distorted 6-s-cis conformation for all these isomers, with the C5,C6,C7,C8 torsional angle for the 7-cis isomers being on average 10° greater than those of analogous 7-trans compounds. The solution conformations of the polyene chain portions of the more important compounds in the 10- and 12-fluororetinal series were investigated by NMR spectroscopic measurements of long-range coupling constants, chemical shifts, and NOE's. The results show that the polyene chains of both 11-cis-10-fluoro- and 11-cis-12-fluororetinal are essentially planar in the regions 7C to 10C and 13C to 15C, but are twisted from planarity around the 10,11 single bonds. Both isomers were found to exist as an equilibrium of 12-s-cis and 12-s-trans conformers. In CDC13 at low temperature, the distorted 12-s-trans conformation appears to be preferred for both isomers. n-Propylamine Schiff bases and protonated Schiff bases derived from some 10- and 12-fluororetinal isomers were prepared as model visual pigment chromophores. These imines and protonated imines were studied in detail using UV-vis and NMR spectroscopy. An improved procedure for the extraction of bovine opsin is presented. 19 F-NMR spectra were recorded for 10- and 12-fluororhodopsin as well as for 9-cis-10-fluororhodopsin. The 19F signals obtained at 15°C were quite broad, with the half-height widths greater than 100 Hz, thus much of the structurally useful information was sacrificed. At this preliminary stage, it is not clear to what extent the dipolar relaxation or chemical shift anisotropy contribute to the observed line-broadening.