ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09706.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09707.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Abstract—Four human cone pigment apoproteins were expressed by transfection of human tissue culture cells with the corresponding complementary DNA clones. Following reconstitution of the cone pigments by incubation with 11‐cisretinal, photobleaching difference absorption spectra were obtained for the blue pigment, the green pigment, and two polymorphic variants of the red pigment. These spectra were analyzed to determine the wavelengths of maximal absorbance and the bandwidths. The recombinant cone pigment spectra were compared to human cone spectral sensitivities and cone pigment absorption spectra determined by microspectrophotometry, single‐cell electrophysiology, reflection densitometry, electroretinography, and psychophysical color and brightness mat

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09708.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Abstract—An attempt has been made to construct a 3‐D model of rhodopsin, a member of G‐protein coupled receptors. Sequence homology of rhodopsin with the latter was a factor considered in the modeling procedure. The constructed model has been used to compare currently available specific protein/substrate interaction information, the shape of the binding cavity derived from shape of binding retinal isomers and analogs and challenged to explain recently available results from a series of fluorinated rhodo

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09709.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Abstract—Bacteriorhodopsin (bR), a light‐driven proton pump, consists of a bundle of seven membrane‐spanning alpha‐helices connected to each other by short extramembranous loops. Previously it has been shown that bR can be reconstituted from three fragments corresponding to the first helix, the second helix, and the remaining five helices, and that this reconstituted material reforms the native structure of bR. In this study, it is shown that the native function is also recovered. Low‐temperature spectroscopy was used to examine the photochemical properties of bR reconstituted from three fragments. At room temperature at pH 6, the reconstituted material shows essentially the same absorption spectrum as native bR, while upon raising the pH at room temperature or cooling the sample in glycerol, a second, blue‐shifted peak appears. The pH and temperature dependence of the absorption spectrum indicates that the reconstituted bR is in an equilibrium between two pigments, which we call P560 and P480. Both pigments convert to their own K intermediates, which differ in absorption maxima, upon illumination with green light at —180°C. Each K intermediate can be reverted to its initial state by light. Similarly, both pigments convert to their own M intermediates upon irradiation with yellow light at ‐77°C. The M intermediate of both species can be reverted only to P560 by light. Both pigments are therefore photoactive. These unique photochemical properties of bR reconstituted from three fragments may be attributable to the lack of a covalent linkage in the loop connecting the A and B helices, and thus possibly to a change in the orientati

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09710.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Abstract—In this paper a consecutive synthetic scheme is described that allows the synthesis of multiply13C labeled retinals with labels in the conjugated chain. This scheme can be extended to incorporation of2H labels in the conjugated chain. Also carotenoids can be multiply13C labeled with this strategy. The same approach has been used to label amino acid

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09711.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09712.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Abstract—For investigation of the chromophore conformation around the trimethyl cyclohexene ring and of the origin of the induced β‐circular dichroism band in rhodopsin, two C6‐C7single bond‐fixed retinal analogues, 6s‐cb‐ and 6s‐trans‐locked bicyclic retinals (6 and 7, respectively) were synthesized and incorporated into bovine opsin in CHAPS‐PC mixture. 6s‐cb‐ and 6s‐tram‐Locked rhodopsin analogues (8 and9) with A max at 539 and 545 nm, respectively, were formed. Interestingly, both 8 and 9 displayed α‐ and β‐circular dichroism bands. The ellipticity of α‐bands are similar in each other, while the β‐band of 8 was about three times stronger than that of 9. Irradiation of 6s‐trans‐locked rhodopsin, 9, in the presence of hyroxylamine, resulted in the formation of only one of the enantiomers of6s‐rrans‐lockedretinal oxime showing a positive circular dichroism signal at around 390 nm. This fact strongly suggests that the retinal binding site of rhodopsin shows a chiral discrimination. From these experimental results, the interactions between the trimethyl cyclohexene ring portion in the chromophore and the neighbouring protein

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09713.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Abstract—The artificial pigments of bacteriorhodopsin (bR) were synthesized with retinal analogues which have6‐s‐cisfixed conformation. In order to study the shape of binding pocket around β‐ionone ring of retinal, we used three different 6‐s‐cis fixed retinals (1–3), which have a gem‐dimethyl group at a different position of the ring structure. These three 6‐s‐cis fixed retinals produced bR analogue pigments with absorption maxima at 574, 594 and 584 nm, respectively. These analogue pigments showed proton pumping activities upon incorporation into vesicles, indicating that6‐s‐transstructure of retinal is not necessary to pump protons. Although these retinal analogues possess the same conjugated π‐electron systems, the wavelengths of maximum absorbance of the analogue pigments were different from each other, indicating a distinctly steric interaction between retinal ring portion and apoprotein. The analogue pigments were also different from each other in regeneration rate and the number of the final products. These differences resulted from a difference in the position of the dimethyl substituent at the ring structure. Based on the results the molecular structure around β‐ionone ring of retina

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09714.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

Abstract—Halobacterium halobium, grown in a defined medium where tyrosine had been largely replaced with m‐fluorotyrosine, biosynthetically produced purple membrane. Analysis of this membrane by high pressure liquid chromatography of phenylthiocarbamyl derivatized amino acids of membrane acid hydrolysates revealed that up to 50% of the tyrosine was present as the m‐fluorotyrosine form. Yields of the purple membrane decreased as the level of incorporation increased. The experimental purple membrane showed a single19F NMR resonance at ‐61.983 ppm (relative to trifluoroacetic acid). The bacteriorhodopsin (bR) in the purple membrane was normal as assayed by gel electrophor‐esis, isoelectric focusing, circular dichroic spectra, and UV‐visible spectra. However, the fluorinated tyrosine bacteriorhodopsins at near neutral pH exhibited slightly slower rates of proton uptake and a slower M‐state decay with biphasic kinetics reminiscent of alkaline solutions of bR (pH>9). These results imply that the tyrosines in bacteriorhodopsin may play a role in the photoactivated proton translocation process of

ISSN:0031-8655    

DOI:10.1111/j.1751-1097.1992.tb09715.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY