Reconstitution of Gloeobacter violaceus rhodopsin with a light-harvesting carotenoid antenna

Biochemistry. 2009 Nov 24;48(46):10948-55. doi: 10.1021/bi901552x.


We show that salinixanthin, the light-harvesting carotenoid antenna of xanthorhodopsin, can be reconstituted into the retinal protein from Gloeobacter violaceus expressed in Escherichia coli. Reconstitution of gloeobacter rhodopsin with the carotenoid is accompanied by characteristic absorption changes and the appearance of CD bands similar to those observed for xanthorhodopsin that indicate immobilization and twist of the carotenoid in the binding site. As in xanthorhodopsin, the carotenoid functions as a light-harvesting antenna. The excitation spectrum for retinal fluorescence emission shows that ca. 36% of the energy absorbed by the carotenoid is transferred to the retinal. From excitation anisotropy, we calculate the angle between the two chromophores as being ca. 50 degrees , similar to that in xanthorhodopsin. The results indicate that gloeobacter rhodopsin binds salinixanthin in a manner similar to that of xanthorhodopsin and suggest that it might bind a carotenoid also in vivo. In the crystallographic structure of xanthorhodopsin, the conjugated chain of the carotenoid lies on the surface of helices E and F, and the 4-keto ring is immersed in the protein at van der Waals distance from the ionone ring of the retinal. The 4-keto ring is in the space occupied by a tryptophan in bacteriorhodopsin, which is replaced by the smaller glycine in xanthorhodopsin and gloeobacter rhodopsin. Specific binding of the carotenoid and its light-harvesting function are eliminated by a single mutation of the gloeobacter protein that replaces this glycine with a tryptophan. This indicates that the 4-keto ring is critically involved in carotenoid binding and suggests that a number of other recently identified retinal proteins, from a diverse group of organisms, could also contain carotenoid antenna since they carry the homologous glycine near the retinal.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Substitution / genetics
  • Bacteroidetes / chemistry
  • Binding Sites / genetics
  • Carotenoids / chemistry
  • Carotenoids / metabolism*
  • Circular Dichroism
  • Cyanobacteria / genetics
  • Fluorescence Polarization
  • Glycosides / chemistry
  • Glycosides / metabolism*
  • Hydroxylamine / chemistry
  • Molecular Conformation
  • Protein Binding / physiology
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Retinaldehyde / chemistry
  • Rhodopsins, Microbial / chemistry*
  • Rhodopsins, Microbial / genetics
  • Rhodopsins, Microbial / metabolism*
  • Schiff Bases / chemistry
  • Spectrometry, Fluorescence
  • Spectrophotometry


  • Glycosides
  • Recombinant Proteins
  • Rhodopsins, Microbial
  • Schiff Bases
  • salinixanthin
  • Hydroxylamine
  • Carotenoids
  • Retinaldehyde