Abstract
Microbial rhodopsins, many variants of which were developed via genetic engineering, have been widely utilized as optogenetic tools. Understanding the molecular mechanisms is important for designing such tools more efficiently. The dynamics of these proteins upon photoactivation can be studied by light-induced difference Fourier transform infrared (FTIR) spectroscopy. As the structural information involves hydrogen, which is not readily accessible via X-ray crystallography, light-induced difference FTIR spectroscopy is a powerful tool to study the molecular mechanisms of these light-receptive proteins. Low-temperature and time-resolved FTIR spectroscopy on two major microbial rhodopsins—bacteriorhodopsin (BR) and halorhodopsin (HR)—are summarized in this review. The low-temperature method stabilizes the intermediate states by decreasing temperature, whereas the time-resolved method allows direct observation at physiological temperature by using a measurement time short enough for their detection. By measuring the difference spectra in X–H and X–D stretching regions, changes in the hydrogen-bonding networks, including water molecules, were elucidated. It was demonstrated that water molecules played an important role in proton and ion translocation in BR and HR. Therefore, for develop optogenetic tools with favorable molecular properties, it may be important to design protein structures in the presence of internal water molecules.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Bamann C, Kirsch T, Nagel G et al (2008) Spectral characteristics of the photocycle of channelrhodopsin-2 and its implication for channel function. J Mol Biol 375(3):686–694. doi:10.1016/j.jmb.2007.10.072
Barth A (2000) The infrared absorption of amino acid side chains. Prog Biophys Mol Biol 74(3–5):141–173. doi:10.1016/S0079-6107(00), 00021-3 [pii]
Barth A, Zscherp C (2002) What vibrations tell us about proteins. Q Rev Biophys 35(4):369–430. doi:10.1017/S0033583502003815
Beja O, Aravind L, Koonin EV et al (2000) Bacterial rhodopsin: evidence for a new type of phototrophy in the sea. Science 289(5486):1902–1906
Beja O, Spudich EN, Spudich JL et al (2001) Proteorhodopsin phototrophy in the ocean. Nature 411(6839):786–789. doi:10.1038/35081051
Bernstein JG, Boyden ES (2011) Optogenetic tools for analyzing the neural circuits of behavior. Trends Cogn Sci 15(12):592–600. doi:10.1016/j.tics.2011.10.003, 00216-6 [pii]
Boyden ES (2011) A history of optogenetics: the development of tools for controlling brain circuits with light. Biol Rep 3:11. doi:10.3410/B3-11, 11 [pii]
Braiman MS, Bousche O, Rothschild KJ (1991) Protein dynamics in the bacteriorhodopsin photocycle: submillisecond Fourier transform infrared spectra of the L, M, and N photointermediates. Proc Natl Acad Sci U S A 88(6):2388–2392
Deisseroth K (2011) Optogenetics. Nat Methods 8(1):26–29. doi:10.1038/nmeth.f.324, nmeth.f.324 [pii]
Efremov R, Gordeliy VI, Heberle J et al (2006) Time-resolved microspectroscopy on a single crystal of bacteriorhodopsin reveals lattice-induced differences in the photocycle kinetics. Biophys J 91(4):1441–1451. doi:10.1529/biophysj.106.083345
Eisenhauer K, Kuhne J, Ritter E et al (2012) In channelrhodopsin-2 Glu-90 is crucial for ion selectivity and is deprotonated during the photocycle. J Biol Chem 287(9):6904–6911. doi:10.1074/jbc.M111.327700
Fenno L, Yizhar O, Deisseroth K (2011) The development and application of optogenetics. Annu Rev Neurosci 34:389–412. doi:10.1146/annurev-neuro-061010-113817
Furutani Y, Kandori H (2014) Hydrogen-bonding changes of internal water molecules upon the actions of microbial rhodopsins studied by FTIR spectroscopy. Biochim Biophys Acta 1837(5):598–605. doi:10.1016/j.bbabio.2013.09.004
Furutani Y, Shibata M, Kandori H (2005) Strongly hydrogen-bonded water molecules in the Schiff base region of rhodopsins. Photochem Photobiol Sci 4(9):661–666. doi:10.1039/b416698a
Furutani Y, Sudo Y, Kandori H (2008) FTIR studies of protein-protein interaction changes between pharaonis phoborhodopsin and its cognate transducer protein. Curr Top Biochem Res 10(2):63–77
Furutani Y, Fujiwara K, Kimura T et al (2012) Dynamics of dangling bonds of water molecules in pharaonis halorhodopsin during chloride ion transportation. J Phys Chem Lett 3(20):2964–2969. doi:10.1021/Jz301287n
Garczarek F, Gerwert K (2006) Functional waters in intraprotein proton transfer monitored by FTIR difference spectroscopy. Nature 439(7072):109–112. doi:10.1038/nature04231, nature04231 [pii]
Gerwert K (1999) Molecular reaction mechanisms of proteins monitored by time-resolved FTIR-spectroscopy. Biol Chem 380(7–8):931–935. doi:10.1515/Bc.1999.115
Gmelin W, Zeth K, Efremov R et al (2007) The crystal structure of the L1 intermediate of halorhodopsin at 1.9 angstroms resolution. Photochem Photobiol 83(2):369–377. doi:10.1562/2006-06-23-RA-947
Hayashi S, Tajkhorshid E, Kandori H et al (2004) Role of hydrogen-bond network in energy storage of bacteriorhodopsin’s light-driven proton pump revealed by ab initio normal-mode analysis. J Am Chem Soc 126(34):10516–10517. doi:10.1021/Ja047506s
Heberle J (2000) Proton transfer reactions across bacteriorhodopsin and along the membrane. Biochim Biophys Acta 1458(1):135–147, 00064-5 [pii]
Inaguma A, Tsukamoto H, Kato HE et al (2015) Chimeras of channelrhodopsin-1 and -2 from Chlamydomonas reinhardtii exhibit distinctive light-induced structural changes from channelrhodopsin-2. J Biol Chem 290(18):11623–11634. doi:10.1074/jbc.M115.642256
Inoue K, Ono H, Abe-Yoshizumi R et al (2013) A light-driven sodium ion pump in marine bacteria. Nat Commun 4:1678. doi:10.1038/ncomms2689
Ito S, Kato HE, Taniguchi R et al (2014) Water-containing hydrogen-bonding network in the active center of channelrhodopsin. J Am Chem Soc 136(9):3475–3482. doi:10.1021/ja410836g
Kandori H (2000) Role of internal water molecules in bacteriorhodopsin. Biochim Biophys Acta 1460(1):177–191, 00138-9 [pii]
Kandori H, Shichida Y (2000) Direct observation of the bridged water stretching vibrations inside a protein. J Am Chem Soc 122(47):11745–11746. doi:10.1021/Ja0032069
Kato HE, Zhang F, Yizhar O et al (2012) Crystal structure of the channelrhodopsin light-gated cation channel. Nature 482(7385):369–374. doi:10.1038/nature10870
Kolbe M, Besir H, Essen LO et al (2000) Structure of the light-driven chloride pump halorhodopsin at 1.8 A resolution. Science 288(5470):1390–1396, 8554 [pii]
Kouyama T, Kanada S, Takeguchi Y et al (2010) Crystal structure of the light-driven chloride pump halorhodopsin from Natronomonas pharaonis. J Mol Biol 396(3):564–579, doi:S0022-2836(09)01455-7 [pii]
Lakatos M, Groma GI, Ganea C et al (2002) Characterization of the azide-dependent bacteriorhodopsin-like photocycle of salinarum halorhodopsin. Biophys J 82(4):1687–1695. doi:10.1016/S0006-3495(02)75521-5
Lanyi JK (1993) Proton translocation mechanism and energetics in the light-driven pump bacteriorhodopsin. Biochim Biophys Acta 1183(2):241–261
Lanyi JK (2004) X-ray diffraction of bacteriorhodopsin photocycle intermediates. Mol Membr Biol 21(3):143–150. doi:10.1080/09687680410001666345
Lorenz-Fonfria VA, Resler T, Krause N et al (2013) Transient protonation changes in channelrhodopsin-2 and their relevance to channel gating. Proc Natl Acad Sci U S A 110(14):E1273–E1281. doi:10.1073/pnas.1219502110
Luecke H, Schobert B, Richter HT et al (1999) Structure of bacteriorhodopsin at 1.55 A resolution. J Mol Biol 291(4):899–911. doi:10.1006/jmbi.1999.3027, S0022-2836(99)93027-9 [pii]
Maeda A (2001) Internal water molecules as mobile polar groups for light-induced proton translocation in bacteriorhodopsin and rhodopsin as studied by difference FTIR spectroscopy. Biochem Biokhimiia 66(11):1256–1268
Maeda A, Sasaki J, Yamazaki Y et al (1994) Interaction of aspartate-85 with a water molecule and the protonated Schiff base in the L intermediate of bacteriorhodopsin: a Fourier-transform infrared spectroscopic study. Biochemistry 33(7):1713–1717
Nagel G, Ollig D, Fuhrmann M et al (2002) Channelrhodopsin-1: a light-gated proton channel in green algae. Science 296(5577):2395–2398. doi:10.1126/science.1072068
Nagel G, Szellas T, Huhn W et al (2003) Channelrhodopsin-2, a directly light-gated cation-selective membrane channel. Proc Natl Acad Sci U S A 100(24):13940–13945. doi:10.1073/pnas.1936192100
Nakanishi T, Kanada S, Murakami M et al (2013) Large deformation of helix F during the photoreaction cycle of Pharaonis halorhodopsin in complex with azide. Biophys J 104(2):377–385. doi:10.1016/j.bpj.2012.12.018
Nishimura S, Sasaki J, Kandori H et al (1995) Structural changes in the lumirhodopsin-to-metarhodopsin I conversion of air-dried bovine rhodopsin. Biochemistry 34(51):16758–16763
Oesterhelt D, Stoeckenius W (1971) Rhodopsin-like protein from the purple membrane of Halobacterium halobium. Nat New Biol 233(39):149–152
Pebay-Peyroula E, Neutze R, Landau EM (2000) Lipidic cubic phase crystallization of bacteriorhodopsin and cryotrapping of intermediates: towards resolving a revolving photocycle. Biochim Biophys Acta 1460(1):119–132
Pfefferle JM, Maeda A, Sasaki J et al (1991) Fourier transform infrared study of the N intermediate of bacteriorhodopsin. Biochemistry 30(26):6548–6556
Radu I, Bamann C, Nack M et al (2009) Conformational changes of channelrhodopsin-2. J Am Chem Soc 131(21):7313–7319. doi:10.1021/ja8084274
Ritter E, Stehfest K, Berndt A et al (2008) Monitoring light-induced structural changes of Channelrhodopsin-2 by UV-visible and Fourier transform infrared spectroscopy. J Biol Chem 283(50):35033–35041, doi:M806353200 [pii]
Rothschild KJ (1992) FTIR difference spectroscopy of bacteriorhodopsin: toward a molecular model. J Bioenerg Biomembr 24(2):147–167
Rummel G, Hardmeyer A, Widmer C et al (1998) Lipidic cubic phases: new matrices for the three-dimensional crystallization of membrane proteins. J Struct Biol 121(2):82–91. doi:10.1006/jsbi.1997.3952
Salom D, Lodowski DT, Stenkamp RE et al (2006) Crystal structure of a photoactivated deprotonated intermediate of rhodopsin. Proc Natl Acad Sci U S A 103(44):16123–16128. doi:10.1073/pnas.0608022103
Sasaki J, Brown LS, Chon YS et al (1995) Conversion of bacteriorhodopsin into a chloride ion pump. Science 269(5220):73–75
Schotte F, Cho HS, Kaila VR et al (2012) Watching a signaling protein function in real time via 100-ps time-resolved Laue crystallography. Proc Natl Acad Sci U S A 109(47):19256–19261. doi:10.1073/pnas.1210938109
Shibata M, Tanimoto T, Kandori H (2003) Water molecules in the schiff base region of bacteriorhodopsin. J Am Chem Soc 125(44):13312–13313. doi:10.1021/ja037343s
Shibata M, Muneda N, Sasaki T et al (2005) Hydrogen-bonding alterations of the protonated Schiff base and water molecule in the chloride pump of Natronobacterium pharaonis. Biochemistry 44(37):12279–12286. doi:10.1021/bi050726d
Siebert F (1995) Infrared spectroscopy applied to biochemical and biological problems. Method Enzymol 246:501–526
Srajer V, Teng T, Ursby T et al (1996) Photolysis of the carbon monoxide complex of myoglobin: nanosecond time-resolved crystallography. Science 274(5293):1726–1729
Tanimoto T, Furutani Y, Kandori H (2003) Structural changes of water in the Schiff base region of bacteriorhodopsin: proposal of a hydration switch model. Biochemistry 42(8):2300–2306. doi:10.1021/bi026990d
Varo G (2000) Analogies between halorhodopsin and bacteriorhodopsin. Biochim Biophys Acta 1460(1):220–229, doi:S0005-2728(00)00141-9 [pii]
Yoshizawa S, Kumagai Y, Kim H et al (2014) Functional characterization of flavobacteria rhodopsins reveals a unique class of light-driven chloride pump in bacteria. Proc Natl Acad Sci U S A 111(18):6732–6737. doi:10.1073/pnas.1403051111
Zhang F, Wang LP, Brauner M et al (2007) Multimodal fast optical interrogation of neural circuitry. Nature 446(7136):633–639. doi:10.1038/nature05744
Acknowledgments
The author thanks many collaborators, especially Professor Hideki Kandori, for studies in this review article and the research grants from KAKENHI (22770159, 22018030, 21023014, 21026016) and JST PRESTO ‘Chemical conversion of light energy’. The authors would like to thank Enago (www.enago.jp) for the English language review.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Japan
About this chapter
Cite this chapter
Furutani, Y. (2015). Molecular Mechanisms for Ion Transportation of Microbial Rhodopsins Studied by Light-Induced Difference FTIR Spectroscopy. In: Yawo, H., Kandori, H., Koizumi, A. (eds) Optogenetics. Springer, Tokyo. https://doi.org/10.1007/978-4-431-55516-2_5
Download citation
DOI: https://doi.org/10.1007/978-4-431-55516-2_5
Publisher Name: Springer, Tokyo
Print ISBN: 978-4-431-55515-5
Online ISBN: 978-4-431-55516-2
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)