Interaction of 1-Hydroxypyrene with BSA Using Fluorescence Anisotropy and Synchronous Fluorescence Analysis Methods†

doi:10.7503/cjcu20150119

Abstract

Abstract:

Fluorescence anisotropy was employed to investigate the interaction of 1-hydroxypyrene(1-OHP), the metabolism biomarker of PAHs in vivo, with a model transport protein of bovine serum albumin(BSA) under simulated physiological conditions. Combined with synchronous fluorescence spectra, the conformation transition of BSA was also investigated. The experiment results showed that 1-OHP can bind to BSA strongly and a 1∶1 complex was formed, with an average binding equilibrium constant of 3.63×10⁶ L/mol. Also as the amounts of BSA differ, the interaction of 1-OHP and BSA was dominated by strong and weak binding modes alternately. Moreover, 1-OHP can bind to tryptophan residues and induce the conformational and micro-environmental changes of BSA, increasing the tryptophan residue of BSA exposuring to a less hydrophobic micro-environment.

Key words: Fluorescence anisotropy, 1-Hydroxypyrene(1-OHP), Bovine serum albumin(BSA), Binding constant, Conformation change, Interaction mode(Ed.: N, K)

CLC Number:

O657
O625

TrendMD:

ZHANG Jing, CHEN Weixiao, ZHANG Wei, DUAN Ying, ZHU Yuxiu, ZHU Yaxian, ZHANG Yong. Interaction of 1-Hydroxypyrene with BSA Using Fluorescence Anisotropy and Synchronous Fluorescence Analysis Methods^†[J]. Chem. J. Chinese Universities, 2015, 36(8): 1511.

Figures/Tables 5

Fig.1 Excitation(a) and emission(b) spectra of 1-OHP in Tris-HCl buffer solution(pH=7.40)a. λem=386 nm, b. λex=346 nm;c(1-OHP)=5.00×10-6 mol/L.

Fig.2 Relationship between fluorescence anisotropy values of 1-OHP and the concentrations of BSA in Tris-HCl buffer solution(pH=7.40)c(1-OHP)=5.00×10-6 mol/L, 105c(BSA)/(mol·L-1)=0, 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.80, 1.00, 1.20, 1.40, 1.60, 1.80, 2.00. λex =346 nm, λem=386 nm.

Table 1 Values of fB, n and K in different 1-OHP-BSA systems(pH=7.40, 291 K)*

10⁵c(BSA)/ (mol·L^-1)	f_B	n	10^-6 K/ (L·mol^-1)	10⁵c(BSA)/ (mol·L^-1)	f_B	n	10^-6 K/ (L·mol^-1)
0	0			0.60	0.83	0.69	2.74
0.20	0.38	0.95	5.20	0.80	0.89	0.56	2.17
0.25	0.48	0.96	8.94	1.00	0.88	0.44	1.35
0.30	0.54	0.90	3.67	1.20	0.93	0.39	1.85
0.35	0.63	0.90	5.34	1.40	0.97	0.35	3.93
0.40	0.70	0.87	4.47	1.60	0.97	0.31	2.48
0.45	0.74	0.83	3.67	1.80	0.96	0.27	1.79
0.50	0.78	0.78	3.20	2.00	1.00	0.25

Fig.3 Energy transfer efficiency(E)and binding distance(r)of BSA and 1-OHP at different concentration ratios of BSA to 1-OHP in 0.05 mol/L Tris-HCl buffer solution(pH=7.40) at 291 Kc(BSA)/c(1-OHP)=0.50, 0.63, 0.83, 1.25, 2.50, 5.00.

Fig.4 Synchronous fluorescence spectra of BSA in 0.05 mol/L Tris-HCl buffer solution(pH=7.40)at 291 K c(BSA)=5.00×10-6 mol/L; 105c(1-OHP)/(mol·L-1): a—k. 0, 0.10, 0.20, 0.30, 0.40, 0.50, 0.60, 0.70,0.80, 0.90, 1.00; (A) Δλ=15 nm; (B) Δλ=60 nm.

References 33

[1]	Kragh-Hansen U., Dan. Med. Bull., 1990, 37(1), 57—84
[2]	Kim K. H., Jahan S. A., Kabir E., Brown R. J. C., Environ. Int., 2013, 60, 71—80
[3]	Ravindra K., Sokhi R., Van-Grieken R., Atmos. Environ., 2008, 42(13), 2895—2921
[4]	Boysen G., Hecht S. S., Mutat. Res./Rev. Mutat., 2003, 543(1), 17—30
[5]	Skupińska K., Zylm M., Misiewicz I., Kasprzycka-Guttman T., Acta Biochim. Pol., 2006, 53(1), 101—112
[6]	Xu C., Gu J., Ma X., Dong T., Meng X., Spectrochim. Acta A, 2014, 125, 391—395
[7]	Marszalek M., Konarska A., Szajdzinska-Pietek E., Wolszczak M., J. Phys. Chem. B, 2013, 117(50), 15987—15993
[8]	ϕstergaard J., Heegaard N. H. H., Electrophoresis, 2003, 24(17), 2903—2913
[9]	Vuignier K., Guillarme D., Veuthey J. L., Carrupt P. A., Schappler J., Pharmaceut. Biomed., 2013, 74, 205—212
[10]	Wu D., Yan J., Wang J., Wang Q., Li H., Food Chem., 2015, 170, 423—429
[11]	Hovius R.,Vallotton P., Wohland T., Vogel H., Trends Pharmacolo. Sci., 2000, 21(7), 266—273
[12]	Du C. R., Luo X., Wei J. R., He T. T., Zheng X. Y., Lin C. W., Chem. Res. Chinese Universities, 2013, 29(5), 854—860
[13]	Li N., Wei Y. J., Journal of Hebei Normal University(Natural Science Edition), 2003, 27(2), 176—180
	(李娜, 魏永巨. 河北师范大学学报(自然科学版), 2003, 27(2), 176—180)
[14]	Zhang D. P., Lu M. L., Wang H. L., J. Am. Chem. Soc., 2011, 133(24), 9188—9191
[15]	Fuentealba D., Kato H., Nishijima M., Fukuhara G., Mori T., Inoue Y., Bohne C., J. Am. Chem. Soc., 2012, 135(1), 203—209
[16]	Li D. H., Peng X. Y., Ye D., Xu J. G., Chem. J. Chinese. Universities, 1999, 20(8), 1220—1222
	(李东辉, 彭兴跃, 叶东, 许金钩. 高等学校化学学报, 1999, 20(8), 1218—1220)
[17]	Lakowicz J.R., Principles of Fluorescence Spectroscopy, 3rd Edition, Springer US, New York, 2006, 353—382
[18]	Carmona N. A., Cohen B., Organero J. A., Douhal A., J. Photochem. Photobiol. A: Chem., 2012, 234, 3—11
[19]	Ouyang Y. F., Wang Y. S., Li G. Y., Physical Testing and Chemical Analysis Part B: Chemical Analysis, 2011, 47(3), 253—256
	(欧阳运富, 王永生, 李贵英. 理化检验: 化学分册, 2011, 47(3), 253—256)
[20]	Zhang Y., Zhang G. Z., Wang Y. M., Lu J. X., J. Anal. Sci., 2000, 16(6), 445—449
	(张勇, 张贵珠, 王月梅, 卢继新. 分析科学学报, 2000, 16(6), 445—449)
[21]	De S., Girigoswami A., Das S., J. Colloid Interf. Sci., 2005, 285(2), 562—573
[22]	Fabriciova G., Sanchez-Cortes S., Garcia-Ramos J. V., Miskovsky P., J. Raman Spectrosc., 2004, 35(5), 384—389
[23]	Jarvis I. W., Dreij K., Mattsson Å., Jernström B., Stenius U., Toxicology, 2014, 321, 27—39
[24]	Ed.: Förster T., Sinanoglu O., Delocalized Excitation and Excitation Transfer, Modern Quantum Chemistry, Part Ⅲ, Academic Press, New York, 1965, (3), 93—137
[25]	Guo M., Li M. H., Yu Q. S., J. Anal. Sci., 2007, 23(4), 405—409
	(郭明, 李铭慧, 俞庆森. 分析科学学报, 2007, 23(4), 405—409)
[26]	Wang N., Ye L., Zhao B. Q., Yu J. X., Braz. J. Med. Biol. Res., 2008, 41(7), 589—595
[27]	Sułkowska A., J. Mol. Struct., 2002, 614(1), 227—232
[28]	Brunmark P., Harriman S., Skipper P. L., Wishnok J. S., Amin S., Tannenbaum S. R., Chem. Res. Toxicol., 1997, 10(8), 880—886
[29]	Sato H., Chuang V. T. G., Yamasaki K., Yamaotsu N., Watanabe H., Nagumo K., Anraku M., Kadowaki D., Ishima Y., Hirono S., Otagiri M., Maruyama T., PLoS One, 2014, 9(2), e87919
[30]	Zsila F., Mol. Pharm., 2013, 10(5), 1668—1682
[31]	Zhang J. X., Yin Z. N., Wu W., Wand Z. X., He R., Wu Z.X., Chem. Res. Chinese Universities, 2012, 28(6), 963—970
[32]	Militello V., Casarino C., Emanuele A., Giostra A., Pullara F., Leone M., Biophys. Chem., 2004, 107(2), 175—187
[33]	Cao X. M., Li J., Yang X., Duan Y., Wang C. X., Thermochim. Acta, 2008, 467(1), 99—106