Korean Journal of Microbiology (미생물학회지)

The Microbiological Society of Korea (한국미생물학회)
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Photochemical/Biophysical Properties of Proteorhodopsin and Anabaena Sensory Rhodopsin in Various Physical Environments

막 단백질인 Proteorhodopsin과 Anabaena Sensory Rhodopsin의 다양한 측정 환경에 따른 광화학/생물리학적 특성

  • Choi, Ah-Reum (Department of Life Science and Institute of Biological Interfaces, Sogang University) ;
  • Han, Song-I (Department of Life Science and Institute of Biological Interfaces, Sogang University) ;
  • Chung, Young-Ho (Division of Life Sciences, Korea Basic Science Institute) ;
  • Jung, Kwang-Hwan (Department of Life Science and Institute of Biological Interfaces, Sogang University)
  • Received : 2011.03.14
  • Accepted : 2011.03.28
  • Published : 2011.03.31
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Abstract

Rhodopsin is a membrane protein with seven transmembrane region which contains a retinal as its chromophore. Although there have been recently reports on various photo-biochemical features of rhodopsins by a wide range of purifying and measurement methods, there was no actual comparison related to the difference of biochemical characteristics according to their physical environment of rhodopsins. First, proteorhodopsin (PR) was found in marine proteobacteria whose function is known for pumping proton using light energy. Second one is Anabaena sensory rhodopsin (Nostoc sp.) PCC7120 (ASR) which belongs to eubacteria acts as sensory regulator since it is co-expressed with transducer 14 kDa in an operon. In this study, we applied two types of rhodopsins (PR and ASR) to various environmental conditions such as in Escherichia coli membranes, membrane in acrylamide gel, in DDM (n-dodecyl-${\beta}$-D-maltopyranoside), OG (octyl-${\beta}$-D-glucopyranoside), and reconstituted with DOPC (1,2-didecanoyl-sn-glycero-3-phosphocholine). According to the light-induced difference spectroscopy, rhodopsins in 0.02% DDM clearly showed photointermediates like M, and O states which respond to the different wavelengths, respectively and showed the best signal/noise ratio. The laser-induced difference spectra showed the fast formation and decay rate of photointermediates in the DDM solubilized samples than gel encapsulated rhodopsin. Each of rhodopsins seemed to be adapted to its surrounding environment.

Chromophore로 레티날(vitamin A)을 사용하는 막 단백질인 로돕신은 7개의 막 단백질로 이루어진다. 최근 광화학/생물리학 측정 방법이 다양해지면서, 그에 따른 새로운 특성도 함께 다양하게 보고되고 있다. 하지만, 다양한 측정 환경에 따른 그 광화학/생물리학 특성의 차이점에 대한 비교연구는 없는 상태이다. 첫째, 빛 에너지를 이용하여 수소 이온 펌프 역할 하는 것으로 잘 알려져 있는 roteorhodopsin (PR)은 해양 proteobacteria에서 발견 되었다. 둘째, 한 오페론에서 14 kDa 전달자와 같이 발현되면서 신호 전달 기능을 하는 Anabaena sensory rhodopsin (ASR)이 있다. 이에 본 연구에서는 이 두 미생물 로돕신을 이용하여 다양한 조건에서 그 차이를 살펴보았다. 각 단백질이 막에 끼어 있는 상태(membrane state), polyacrylamide gel에 고정되어 있는 상태, nonionic detergent일종인 DDM (n-dodecyl-${\beta}$-D-maltopyranoside)과, OG (octyl-${\beta}$-D-glucopyranoside)에 녹아 있는 상태, 좀더 자연상태와 유사한 환경을 위해 단백질만 깨끗하게 정제한 다음 다시 Escherichia coli의 지질인 DOPC (1,2-didecanoyl-sn-glycero-3-phosphocholine)에 재조립한 상태, 이렇게 5가지 조건에서 각각의 광화학/생물리학 특징을 흡수스펙트럼(absorption spectrum), 빛-어둠 차이 흡수스펙트럼(light-induced difference spectrum), 포토사이클(photocycle)을 측정하였다. 그 결과 DDM에 녹아 있는 PR과 ASR에서 각 다른 파장에서 M과 O state와 같은 선명한 photointermediate를 가지고, 가장 signal/noise 비율이 좋았다. 본 연구를 통해 막단백질의 다양한 측정 환경에 대한 그 특성의 차이점을 살펴봄으로써 앞으로 다양한 종류의 로돕신 연구에 기초 기반이 될 것으로 사료된다.

Keywords

References

  1. Beja, O., E.N. Spudich, J.L. Spudich, M. Leclerc, and E.F. DeLong. 2001. Proteorhodopsin phototrophy in the ocean. Nature 411, 786-789. https://doi.org/10.1038/35081051
  2. Bogomolni, R.A. and J.L. Spudich. 1982. Identification of a third rhodopsin-like pigment in photoactic Halobacterium halobium. Proc. Natl. Acad. Sci. USA 79, 4308-4312. https://doi.org/10.1073/pnas.79.14.4308
  3. Brown, L.S. 2004. Fungal rhodopsins and opsin-related proteins: eukaryotic homologues of bacteriorhodopsin with unknown functions. Photochem. Photobiol. Sci. 3, 555-565. https://doi.org/10.1039/b315527g
  4. Choi, A.R., S.Y. Kim, S.R. Yoon, K. Bae, and K.H. Jung. 2007. Substitution of Pro206 and Ser86 residues in the retinal binding pocket of Anabaena sensory rhodopsin is not sufficient for proton pumping function. J. Microbiol. Biotechnol. 17, 138-145.
  5. Choi, A.R., S.Y. Kim, S.R. Yoon, and K.H. Jung. 2005. Glu-56 in HtrI is critical for phototaxis signaling in Halobacterium salinarum. Intergrative Biosciences 9, 139-144. https://doi.org/10.1080/17386357.2005.9647264
  6. Jung, J.Y., A.R. Choi, Y.K. Lee, H.K. Lee, and K.H. Jung. 2008. Spectroscopic and photochemical analysis of proteorhodopsin variants from the surface of the Arctic ocean. FEBS 582, 1679-1684. https://doi.org/10.1016/j.febslet.2008.04.025
  7. Jung, K.H. 2007. The distinct signaling mechanism of microbial sensory rhodopsins in Archaea, Eubacteria and Eukarya. Photochem. Photobiol. 83, 63-69. https://doi.org/10.1562/2006-03-20-IR-853
  8. Jung, K.H., V.D. Trivedi, and J.L. Spudich. 2003. Demonstration of a sensory rhodopsin in eubacteria. Mol. Microbiol. 47, 1513- 1522. https://doi.org/10.1046/j.1365-2958.2003.03395.x
  9. Kawanabe, A., Y. Furutani, K.H. Jung, and H. Kandori. 2007. Photochromism of Anabaena sensory rhodopsin. J. Am. Chem. Soc. 129, 8644-8649. https://doi.org/10.1021/ja072085a
  10. Kim, S.Y., S.A. Waschuk, L.S. Brown, and K.H. Jung. 2008. Screening and characterization of proteorhodopsin color-tuning mutations in Escherichia coli with endogenous retinal synthesis. BBA. 1777, 504-513. https://doi.org/10.1016/j.bbabio.2008.03.010
  11. Matsuno-Yagi, A. and Y. Mukohata. 1977. Two possible roles of bacteriorhodopsin: a comparative study of strains of Halobacterium halobium differing in pigmentation. Biochem. Biophys. Res. Comm. 78, 237-243. https://doi.org/10.1016/0006-291X(77)91245-1
  12. Mylene, M.R.M., A.R. Choi, L. Shi, A.G. Bezerra, Jr., K.H. Jung, and L.S. Brown. 2009. The photocycle and proton translocation pathway in a cyanobacterial ion-pumping rhodopsin. Biophys. J. 96, 1471-1481. https://doi.org/10.1016/j.bpj.2008.11.026
  13. Oesterhelt, D. and W. Stoechenius. 1973. Functions of a new photoreceptor membrane. Proc. Natl. Acad. Sci. USA 70, 2853- 2857. https://doi.org/10.1073/pnas.70.10.2853
  14. Ovichinnilov, YuA. 1982. Rhodopsin and bacteriorhodopsin: structure-function relations. FEBS Lett. 148, 179-191. https://doi.org/10.1016/0014-5793(82)80805-3
  15. Sehobert, B. and J.K. Lanyi. 1982. Halorhodopsin is a lightdriven chloride pump. J. Biol. Chem. 257, 10306-10313.
  16. Shi, L., S.R. Yoon, A.G. Bezerra Jr., K.H. Jung, and L.S. Brown. 2006. Cytoplasmic shuttling of protons in anabaena sensory rhodopsin: implications for signaling mechanism. J. Mol. Biol. 358, 686-700. https://doi.org/10.1016/j.jmb.2006.02.036
  17. Spudich, E.N. and J.L. Spudich. 1982. Control of transmembrane ion fluxes to select halorhodopsin-deficient and other energytransduction mutants of Halobacterium halobium. Proc. Natl. Acad. Sci. USA 79, 4308-4312. https://doi.org/10.1073/pnas.79.14.4308
  18. Spudich, J.L. and K.H. Jung. 2005. Microbial rhodopsin: phylogenetic and functional diversity, Handbook of Photosensory Receptros, pp. 1-24. In W.R. Briggs and J.L. Spudich (eds.), Wiley-VCH.
  19. Spudich, J.L., C.S. Yang, K.H. Jung, and E.N. Spudich. 2000. Retinylidene proteins: structures and functions from archaea to humans. Annu. Rev. Cell Dev. Biol. 16, 365-392. https://doi.org/10.1146/annurev.cellbio.16.1.365
  20. Takahashi, T., Y. Mochizuki, N. Kamo, and Y. Kobatake. 1985. Evidence that the long-lifetime photointermediate of s-rhodopsin is a receptor for negative phototaxis in Halobacterium halobium. Biochem. Biophys. Res. Commun. 127, 99-105. https://doi.org/10.1016/S0006-291X(85)80131-5