吾人以時間解析傅氏紅外光譜儀研究細菌視紫質的光迴圈行為，並由紅外差異吸收光譜中挑選出12支特徵吸收峰，將其時間側寫以相關性方法進行分析，繪製出數個時間區段內的相關性等高線圖。藉由觀察等高線圖之中相關性反轉，分析光迴圈中的動力學行為受到驅動力影響的強度及時序，其結果與前人對光迴圈中間態演進的研究吻合。吾人希望提供一個不需要建立中間態以及整體指數擬合分析的方法，直接由吸收峰之間隨時間變化的關聯性，獲得光迴圈中質子傳遞過程的動力學資訊。

A step-scan Fourier-transform interferometer was employed to collect the time-evolved difference infrared spectra of bacteriorhodopsin upon photoexcitation. Without a kinetics model being established or target analysis to extract the difference spectra of the intermediates in the conventional photocycle, correlation analysis of the infrared features was used to examine the evolution of the interactive strengths between the retinal Schiff base and its proton donor and acceptor in different periods of the photocycle. Following the evolution of the correlation, those correlative characteristics satisfactorily coincided with the transitions of M  N, N  O, and O  bR. We therefore have illustrated the trajectory of the driving forces on the localized molecular moieties during the photocycle: proton acceptor Asp85 (1762 and 1755 cm–1) → proton donor Asp96 (1400 cm–1) → C–C stretch of 13-cis retinal (1186 cm–1) → C14=C15 stretch of all-trans retinal (1506 cm–1) → C10–C11 stretch of all-trans retinal (1168 cm–1). The correlation analysis successfully provided the dynamic and local conformational alterations as the bR photocycle evolved, without the need to establish a kinetics model, introduce discrete intermediates, or use deconvolution algebra. Including more bands in the correlation analysis provides more thorough information of the structural alteration of the photocycle. The combination of the time-resolved infrared spectroscopic method and the correlation analysis could be a promising method of illustrating the photochemistry of photosynthetic proteins at the molecular level.

[1] Oesterhelt, D.; Stoeckenius, W. Proc. Nat. Acad. Sci. U. S. A. 1973, 70, 2853−2857.
[2] Racker, E.; Stoeckenius, W. J. Bio. Chem. 1974, 249, 662−663.
[3] Gai, F.; Hasson, K. C.; McDonald, J. C.; Anfinrud, P. A. Science 1998, 279, 1886−1891.
[4] Stoeckenius, W.; Rowen, R. J. Cell Biol. 1967, 34, 365−393.
[5] Stoeckenius, W.; Kunau, W. H. J. Cell Biol. 1968, 38, 337−357.
[6] Oesterhelt, D.; Stoeckenius, W. Nat. New Biol. 1971, 233, 149−152.
[7] Blaurock, A. E.; Stoeckenius, W. Nat. New Biol. 1971, 233, 152−155.
[8] Lozier, R. H.; Bogomolni, R. A.; Stoeckenius, W. Biophys. J. 1975, 15, 955−962.
[9] Terner, J.; Campion, A.; El-Sayed, M. A. Proc. Nat. Acad. Sci. U. S. A. 1977, 74, 5212−5216.
[10] Eisfeld, W.; Pusch, C.; Diller, R.; Lohrmann, R.; Stockburger, M. Biochemistry 1993, 32, 7196−7215.
[11] Olejnik, J.; Brzezinski, B.; Zundelb, G. J. Mol. Structure 1992, 271, 157−173.
[12] Zscherp, C.; Heberle, J. J. Phys. Chem. B 1997, 101, 10542−10547.
[13] Lórenz-Fonfría, V. A.; Kandori, H. J. Am. Chem. Soc. 2009, 131, 5891–5901.
[14] Du, M.; and Fleming, G. R. Biophys. Chem. 1993, 48, 101–111.
[15] Groot, H. J. M.; Harbison, G. S.; Herzfeld, J.; Griffin, R. G. Biochemistry, 1989, 28, 3346–3353.
[16] Mak-Jurkauskas, M. L.; Bajaj, V. S.; Hornstein, M. K.; Belenky, M.; Griffin, R. G; Herzfeld, J. Proc. Nat. Acad. Sci. U. S. A. 2008, 105, 883−888.
[17] K hlbrandt, W. Nature 2000, 406, 569−570.
[18] Luecke, H.; Schobert, B.; Richard, H. T.; Cartailler, J. P.; Lanyi, J. K. J. Mol. Biol. 1999, 291, 899−911.
[19] Baudry, J.; Tajkhorshid, E.; Molnar, F.; Phillips, J.; Schulten, K. J. Phys. Chem. B 2001, 105, 905−918.
[20] Lanyi, J. K. Annu. Rev. Physiol. 2004, 66, 665−688.
[21] Garczarek, F.; Gerwert, K. Nature 2006, 439, 109−112.
[22] Lanyi, J. K. Biochim. Biophys. Acta 2006, 1757, 1012−1018.
[23] Lanyi, J. K.; A Structural View of Proton Transport by Bacteriorhodopsin; Royal Society of Chemistry, 2005.
[24] Henderson, H. J. Mol. Biol. 1975, 93, 123−138
[25] Kate, M.; Kushwaha, S. C.; Sprott, G. D. Methods Enzymol. 1982, 88, 98−111.
[26] Dracheva, S.; Bose, S.; Hendler, R. W. FEBS Lett. 1996, 382, 209−212.
[27] Birge, R. R.; Gillespie, N. B.; Izaguirre, E. W.; Kusnetzow, A.; Lawrence, A. F.; Singh, D.; Wang, S. Q.; Schmidt, E.; Stuart, J. A.; Seetharaman, S.; Wise, K. J. J.
77
Phys. Chem. B 1999, 103, 10746−10766.
[28] Edman, K.; Nollert, P.; Royant, A.; Belrhali, H.; Pebay-Peyroula, E.; Hajdu, J.; Neutze, R.; Landau, E. M. Nature 1999, 401, 822−826.
[29] Brown, L. S.; Lanyi, J. K. Proc. Natl. Acad. Sci. U. S. A. 1996, 93, 1731−1734.
[30] Royant, A.; Edman, K.; Ursby, T.; Peybay-Peyroula, E.; Landau, E. M.; Neutze, R. Nature 2000, 406, 645−648.
[31] Balashov, S. P.; Govindjee, R.; Kono, M.; Imasheva, E.; Lukashev, E.; Ebrey, T. G.; Crouch, R. K.; Menick, D. R.; Feng, Y. Biochemistry 1993, 32, 10331−10343.
[32] Lanyi, J. K.; J. Biol. Chem. 1997, 272, 31209−31212.
[33] Braiman, M. S.; Bousche, O.; Rothschild, K. J. Proc. Natl. Acad. Sci. U. S. A. 1991, 88, 2388−2392.
[34] Smith, S. O.; Pardoen, J. A.; Mulder, P. P. J.; Curry, B.; Lugtenburg, J.; Mathies, R. Biochemistry 1983, 22, 6141−6148.
[35] Morgan, J. E.; Vakkasoglu, A. S.; Gennis, R. B.; Maeda, A. Biochemistry 2007, 46, 2787−2796.
[36] Scherrer, P.; Mathew, M. K.; Sperling, W.; Stoeckenius, W. Biochemistry 1989, 28, 829−834.
[37] Wang, J.; Link, S.; Heyes, C. D.; El-Sayed, M. A. Biophys. J. 2002, 83, 1557−1566.
[38] Renthal, R.; Gracia, N.; Regalado, R. Photochem. Photobiol. 2000, 72, 714−718.
[39] Dioumaev, A. K.; Biochemistry 2001, 66, 1269−1276.
[40] Tamm, L. K.; Tatulian, S. A. Quart. Rev. Biophys. 1997, 30, 365−429.
[41] Dwivedi, A. M.; Krimm, S. Biopolym. 1984, 23, 2025−2065. (HERE)
[42] Krimms, S.; Dwivedi, A. M. Science 1982, 216, 407−408.
[43] Barth, A.; Zscherp, C. Quart. Rev. Biophys. 2002, 35, 369−430.
[44] Barth, A. Progr. Biophys. Mol. Biol. 2000, 74, 141−173.
[45] Barnett, S. M.; Dracheva, S.; Hendler, R. W.; Levin, I. W. Biochemistry 1996, 35, 4558−4567.
[46] Arrondo, J. L. R.; Go I, F. M. Chem. Phys. Lipids 1998, 96, 53−68.
[47] Sasaki, J.; Lanyi, J. K.; Needleman, R.; Yoshizawa, T.; Maeda, A. Biochemistry 1994, 33, 3178−3184.
[48] Rothschild, K. J.; Degrip, W. J.; Sanches, R. Biochem. Biophys. Acta 1980, 596, 338−351.
[49] Rothschild, K, J.; Grip, W. J. D.; Stanley, H. E. Science 1976, 191, 1176−1178.
[50] Ahmed, H. Principles and reactions of protein extraction, purification and characterization; Boca Raton, USA, 2005.
[51] Rothschild, K. J.; Braiman, M. S.; He, Y. W.; Marti, T.; Khorana, H. G. J. Biol. Chem. 1990, 265, 16985−16991.
78
[52] Lórenz-Fonfría, V. A.; Furutani, Y.; Kandori, H. Biochemistry 2008, 47, 4071–4081.
[53] Siebert, F.; Mäntele, W. Eur. J. Biochem. 1983, 130, 565–571.
[54] Rothschild, K. J.; Roepe, P.; Lugtenburg, J.; Pardoen, J. A. Biochemistry 1984, 23, 6103–6109.
[55] Zscherp, C.; Schlesinger, R.; Tittor, J.; Oesterhelt, D.; Heberle, J. Proc. Natl. Acad. Sci. U. S. A. 1999, 96, 5498–5503.
[56] Morgan, J. E.; Vakkasoglu, A. S.; Lugtenburg, J.; Gennis, R. B.; Maeda, A. Biochemistry 2008, 47, 11598–11605.
[57] Wang, J.; El-Sayed, M. A. Biophys. J. 2001, 80, 961–971.
[58] Braiman, M. S.; Mogi, T.; Marti, T.; Stern, L. J.; Khorana, H. G.; Rothschild, K. J. Biochemistry 1988, 27, 8516−8520.
[59] Smith, S. O.; Myers, A. B.; Mathies, R. A.; Pardoen, J. A.; Winkel, C.; Van Den Berg, E. M. M.; Lugtenburg, J. Biophys. J. 1985, 47, 653–664.
[60] Riesle, J.; Oesterhelt, D.; Dencher, N. A.; Heberle, J. Biochemistry 2006, 35, 6635–6643.
[61] Bernath, P. F. Encyclopedia of Analytical Science, 2nd.: Fourier Transform Techniques; Elsevier, 2005.
[62] Wartewig, S. IR and Raman Spectroscopy:Fundamental Processing; Wiley-VCH, 2005.
[63] Kauppinen, J.; Partanen, J. Fourier Transform in Spectroscopy; Wiley-VCH, 2011.
[64] Griffiths, P. R.; Haseth, J. A. Fourier Transform Infrared Spectrometry, 2nd.; Wiley, 2007.
[65] Jiang, E. Y. Advanced FT-IR Spectroscopy; Thermo Electron Corporation, 2003.
[66] Uhmann, W.; Becker, A.; Taran, C.; Siebert, F. Appl. Spectrosc. 1991, 45, 390-397.
[67] Chu, L. K.; Lee, Y. P. Instruments Today 2009, 31, 27−35.
[68] Noguchi, T.; Sugiura, M. Biochemistry 2002, 41, 2322–2330.
[69] Mark, S. B,; Olaf, B.; Kenneth, J. R. Proc. Natl. Acad. Sci. U.S.A. 1991, 88, 2388-2392.
[70] Hartland, G. V.; Qin, D.; Dai, H. L. J. Chem. Phys. 1994, 100, 7832.
[71] Yeh, P. S.; Leu, G. H.; Lee, Y. P.; Chen, I. C. J. Chem. Phys. 1995, 103, 4879.
[72] Hu, X.; Frei, H.; Spiro, T. G. Biochemistry 1996, 35, 13001−13005.
[73] Huang, C. Y.; Getahun, Z.; Wang, T.; DeGrado, W. F.; Gai, F. J. Am. Chem. Soc. 2001, 123, 12111−12112.
[74] Vasenkov, S.; Frei, H. J. Phys. Chem. A 2000, 104, 4327−4332.
[75] Snellenburg, J. J.; Laptenok, S. P.; Seger, R.; Mullen, K. M.; van Stokkum, I. H. M. J. Statist. Software 2012, 49, 1–22.
79
[76] Mertz, L. Transformations in Optics; Wiley, 1965.
[77] Mertz, L. Infrared Phys. 1967, 7, 17–23.
[78] Geerts, Y.; Steyaert, M.; Sansen, W. M. C. Design of Multi-Bit Delta-Sigma A/D Converters; Springer Science & Business Media, 2002.
[79] Fellgett, P. B. J. Phys. Radium 1958, 19, 187−191.
[80] Jacquinot, P. Rep. Progr. Phys. 1960, 23, 267−312.
[81] Connes, J.; Connes, P. J. Opt. Soc. Am. 1966, 56, 896−910.
[82] Robin, R.; Gregor, H.; Marthias, L.; Klaus, G. Biochemistry 1998, 37, 5001−5009.
[83] Christian, Z.; Ramona, S.; Jorg, T.; Dieter, O.; Joachim, H. Proc. Natl. Acad. Sci. U.S.A. 1999, 96, 5498−5503.
[84] Lanyi, J, K., Bacteriorhodopsin, Chemistry of. In Wiley Encyclopedia of Chemical Biology, Begley, T.P., ED. John Wiley & Sons, Inc.: 2007.
[85] Morgan, J. E.; Vakkasoglu, A. S.; Lanyi, J. K.; Lugtenburg, J.; Gennis, R. B.; Maeda, A. Biophys. J. 2012, 103, 444−452.
[86] Tittor, J.; Paula, S.; Subramaniam, S.; Heberle, J.; Henderson, R.; Oesterhelt, O., J. Mol. Biol. 2002, 319, 555−565.