Ectothiorhodospira halophila

Fig. 2. H NMR spectra of (A) oxidized spinach Fe2S2 ferredoxin (33) (B) reduced spinach Fe2S2 ferredoxin (5f) (C) oxidized Desulfovibrio gigas Fe3S4 ferredoxin (138) (D) oxidized ectothiorhodospira halophila HiPIP iso-II (23) (E) reduced Chromatium vinosum HiPIP (14) (F) fully reduced Clostridium pasteurianum 2(Fe4S4) ferredoxin (139). Chemical shift values are in ppm.

Fig. 5. H NMR spectrum of oxidized Ectothiorhodospira halophila HiPIP iso-II (A) and experimental temperature dependence of the shifts of the signals (B) (18).

Ectothiorhodospira halophila HiPIPs, 38 249 Edge studies, photosystem II, 33 228-230 EDTA... 

From such a viewpoint, we are examining primary processes of photoreactions of PYP [1] which functions as a blue light photoreceptor for a negative phototaxis of the purple sulfur bacterium Ectothiorhodospira halophila, some FP s [2] and Rh [3] by means of the fs fluorescence up-conversion measurements. In this article, we will discuss our latest results of fs fluorescence dynamics studies on PYP, because PYP is very stable for repeated irradiation which induces photocycles so that the very accurate experimental results can be obtained rather easily and also the preparation of the site-directed mutants as well as the PYP analogues with modified chromophores are rather easy. However, before that, we will summarize briefly results of our previous investigations. 

W. Sprenger, W.D. Hoff, J.P. Armitage, K. Hellingwerf (1993). The eubacterium Ectothiorhodospira halophila is negatively phototactic, with a wavelength dependence that fits the absorption spectrum of the photoactive yellow protein. J. Bacteriol, 175, 3096-3105. 

Photoactive yellow protein (PYP) was discovered 20 year ago in Halorhodospira halophila, then known as Ectothiorhodospira halophila [1,2]. In several halophilic purple bacteria it has a vital role in the avoidance response to blue light (phototaxis). It has been thoroughly studied as a model photoreceptor system and as the structural prototype for the PAS class of signal transduction proteins. PYP has 125 amino acid residues in an a// -fold with six antiparallel /1-sheets and several helices (see Fig. 5.1). The covalently bound p-coumaric acid chromophore is linked to the only cysteine in the protein (Cys69) (see Fig. 5.1). Hellingwerf has published an excellent review of the photophysical behavior of PYP [1],... 

Most high-potential iron proteins have been isolated from purple photosynthetic bacteria and vary in size from 6 to 10 kDa the tertiary structures of many have been determined in the crystalline and/or solution states, including HiPIPs from C. vinosum (1,49,52,108-110), Ectothiorhodospira halophila 1 48, 102, 111, 112), Ectothiorhodospira... 

Differential Scanning Calorimetry Data for Tyr-12 Mutants of Ectothiorhodospira halophila I HiPIP... 

FIGURE 5.23 The hybrid maps for the differences (A B), the omissions (C D), and the extrapolations (E F) of the X-ray determined electronic densities of the chromophore 4-hydroxycirmamyl (up) of the yellow photo-active protein that contains it (down), as extracted from the photo-tropic bacterium Ectothiorhodospira halophila, respectively after Genick et al. (1997) Bioinformatics Protein Models (2013). 

Kappl R, Cituli S, Luchinat C, Huttennann J. 1999. Probing structural and electronic properties of the oxidized [Fe4S4] cluster of Ectothiorhodospira halophila iso-II high-potential iron-sulfur protein by ENDOR spectroscopy. J Am Chem Soc 121 1925-1935. 

Breiter DR, Meyer TE, Rayment I, Holden HM. 1991. The molecular structure of the high-potential iron-sulfur protein isolated fi om Ectothiorhodospira halophila determined at 2.5 A resolution. JBiol Chem 266 18660-18667. 

Band L, Bertini I, Capozzi F, Carloni P, Ciurli S, Luchinat C, PiccioU M. 1993. The iron-sulfirr cluster in the oxidized high-potential iron protein fi om Ectothiorhodospira halophila. J Am Chem Soc 115 3431-3440. 

The p-coumaric acid chromophore of the photoactive yeUow protein (PYP) of Ectothiorhodospira halophila (now reclassified Holorhodospira halophila, see Chapter 123)... 

The presently known chromophores are 4-hydroxy-cinnamic acid (e.g., in Ectothiorhodospira halophila, now reclassified as Halorhodospira halophila see Chapter 123), carotenoids (e.g., retinal in Halobacterium salinarum, see Chapter 124), pterins and flavins (e.g., in Euglena gracilis, see Chapter 115), and dian-thronic molecules (e.g., stentorin in S. coeruleus and blepharismins in B. japonicum, see Chapter 122). 

In 1985 several ferredoxins and other chromophoric proteins from the halophihc phototrophic bacterium Ectothiorhodospira halophila were isolated. One of the other chromophoric proteins was yellow and was named photoactive yeUow protein in a subsequent study. E. halophila was reclassified in 1996 to its current name Halorhodospira halophila. H. halophila is a unicellular prokaryote, or more specifically, a phototrophic purple sulfur spirillum that deposits sulfur extraceUularly. It was first isolated and classified from salt-encrusted mud taken from the shores of Summer Lake, Lake County, Oregon. Later, it was also isolated from the extremely saline lakes of the Wadi el Natrun in Egypt. Both locations are salt lakes, and H. halophila only thrives in extremely salty environments. 

Meyer, T.E., Isolation and characterization of soluble cytochromes, ferredoxins and other chro-mophoric proteins from the halophilic phototrophic bacterium Ectothiorhodospira halophila, Bio-chim. Biophys. Acta, 806,1, 175-183,1985. 

Meyer, T.E., Tollin, G., Hazzard, J.H., and Cusanovich, M.A., Photoactive yellow protein from the purple phototrophic bacterium, Ectothiorhodospira halophila. Quantum yield of photobleaching and effects of temperature, alcohols, glycerol, and sucrose on kinetics of photobleaching and recovery, Biophys. /., 56, 3, 559—564, 1989. 

Imamoto, Y, Kataoka, M., and Tokunaga, E, Photoreaction cycle of photoactive yeUow protein from Ectothiorhodospira halophila studied by low-temperature spectroscopy. Biochemistry, 35, 45, 14,047-14,053,1996. 

Hoff, W.D., Kwa, S.L.S., van GrondeUe, R, and HeUingwerf, K.J., Low temperature absorbance and fluorescence spectroscopy of the photoactive yeUow protein from Ectothiorhodospira halophila, Photochem. Photobiol, 56, 529-539, 1992. 

Ujj, L, Devanathan, S., Meyer, T.E., Cusanovich, M.A., Tollin, G., and Atkinson, G.H., New photocycle intermediates in the photoactive yellow protein from Ectothiorhodospira halophila picosecond transient absorption spectroscopy, Biophys. J., 75,1, 406-412,1998. 

Hendriks, J., Hoff, W.D., Crielaard, W., and Hellingwerf, K.J., Protonation/deprotonation reactions triggered by photoactivation of photoactive yellow protein from Ectothiorhodospira halophila, J. Biol Chem., 274, 25, 17,655-17,660,1999. 

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