Chemical reactivity and spectroscopy of the thiol ester-linked p-coumaric acid chromophore in the photoactive yellow protein from Ectothiorhodospira halophila.
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Hoff WD, Devreese B, Fokkens R, Nugteren-Roodzant IM, Van Beeumen J, Nibbering N, Hellingwerf KJ
Chemical reactivity and spectroscopy of the thiol ester-linked p-coumaric acid chromophore in the photoactive yellow protein from Ectothiorhodospira halophila.
Biochemistry. 1996 Jan 30;35(4):1274-81.
- PubMed ID
- 8573584 [ View in PubMed]
- Abstract
We have recently identified p-coumaric acid as the chromophore of the photoactive yellow protein (PYP) from the purple sulfur bacterium Ectothiorhodospira halophila, a blue-light photoreceptor with rhodopsin-like photochemistry [Hoff, W. D., Dux, P., Hard, K., Nugteren-Roodzant, I. M., Crielaard, W., Boelens, R., Kaptein, R., Van Beeumen, J., & Hellingwerf, K. J. (1994) Biochemistry 33, 13959-13962]. Here we report on the chemistry of the linkage of this new photoactive cofactor to apoPYP: (i) Analysis of chromophore-peptide conjugates of PYP by high-resolution mass spectrometry unambiguously shows that the p-coumaric acid molecule is bound to Cys 69 via a thiol ester bond. The PYP chromophore is the first cofactor known to be stably thiol ester-linked to its apoprotein. (ii) The chemical reactivity of this thiol ester bond with respect to dithiothreitol, performic acid, and high pH is similar to that of disulfide bridges. These treatments result in the cleavage of the thiol ester bond, concomitant with strong shifts in the UV/vis absorbance band of the chromophore. (iii) The spectral properties of the PYP chromophore under different conditions are related to the structural integrity of the protein, the presence of the thiol ester bond, and the ionization state of the phenolic proton of the chromophore. These results are important for the general problem of spectral tuning in photoreceptor proteins.