THE XANTHOPSINS - A NEW FAMILY OF EUBACTERIAL BLUE-LIGHT PHOTORECEPTORS

Photoactive yellow protein (PYP) is a photoreceptor that has been isol ated from three halophilic phototrophic purple bacteria. The PYP from Ectothiorhodospira halophila BN9626 is the only member for which the s equence has been reported at the DNA level. Here we describe the cloni ng and sequencing of the genes encoding the PYPs from E.halophila SL-1 (type strain) and Rhodospirillum salexigens. The latter protein conta ins, like the E.halophila PYP, the chromophore trans p-coumaric acid, as we show here with high performance capillary zone electrophoresis. Additionally, we present evidence for the presence of a gene encoding a PYP homolog in Rhodobacter sphaeroides, the first genetically well-c haracterized bacterium in which this photoreceptor has been identified . An ORF downstream of the pyp gene from E.halophila encodes an enzyme , which is proposed to be involved in the biosynthesis of the chromoph ore of PYP. The pyp gene from E.halophila was used for heterologous ov erexpression in both Escherichia coli and R.sphaeroides, aimed at the development of a holoPYP overexpression system (an intact PYP, contain ing the p-coumaric acid chromophore and displaying the 446 nm absorban ce band). In both organisms the protein could be detected immunologica lly, but its yellow color was not observed. Molecular genetic construc tion of a histidine-tagged version of PYP led to its 2500-fold overpro duction in E.coli and simplified purification of the heterologously pr oduced apoprotein. HoloPYP could be reconstituted by the addition of p -coumaric anhydride to the histidine-tagged apoPYP (PYP lacking its ch romophore). We propose to call the family of photoactive yellow protei ns the xanthopsins, in analogy with the rhodopsins.