Conformational changes of β-lactoglobulin in sodium bis(2-ethylhexyl) sulfosuccinate reverse micelles -: A fluorescence and CD study

被引:27
作者
Andrade, SM [1 ]
Carvalho, TI [1 ]
Viseu, MI [1 ]
Costa, SMB [1 ]
机构
[1] Univ Tecn Lisboa, Ctr Quim Estrutural, Inst Super Tecn, P-1049001 Lisbon, Portugal
来源
EUROPEAN JOURNAL OF BIOCHEMISTRY | 2004年 / 271卷 / 04期
关键词
beta-lactoglobulin; conformation; quenching; reverse micelles;
D O I
10.1111/j.1432-1033.2004.03977.x
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The effect of beta-lactoglobulin encapsulation in sodium bis(2-ethylhexyl) sulfosuccinate reverse micelles on the environment of protein and on Trp was analysed at different water contents (omega(0)). CD data underlined the distortion of the beta-sheet and a less constrained tertiary structure as the omega(0) increased, in agreement with a concomitant red shift and a decrease in the signal intensity obtained in steady-state fluorescence measurements. Fluorescence lifetimes, evaluated by biexponential analysis, were tau(1) = 1.28 ns and tau(2) = 3.36 ns in neutral water. In reverse micelles, decay-associated spectra indicated the occurrence of important environmental changes associated with omega(0). Bimolecular fluorescence quenching by CCl4 and acrylamide was employed to analyse alterations in the accessibility of the two Trp residues in beta-lactoglobulin, induced by changes in omega(0). The average bimolecular quenching constant <k(q)(CCl4) was found not to depend on omega(0), confirming the insolubility of this quencher in the aqueous interface, while <k(q)(acrylamide)> increases with omega(0). The drastic decrease with omega(0) of k(q), associated with the longest lifetime, k(q2)(CCl4), comparatively to the increase of <k(q2)(acrylamide), emphasizes the location of beta-lactoglobulin in the aqueous interfacial region especially at omega(0) greater than or equal to 10. The fact that k(q2)(acrylamide) (omega(0) = 30) >> k(q2)(acrylamide) (water) also confirms the important conformational changes of encapsulated beta-lactoglobulin.
引用
收藏
页码:734 / 744
页数:11
相关论文
共 68 条
[1]   INTERPRETATION OF FLUORESCENCE DECAYS IN PROTEINS USING CONTINUOUS LIFETIME DISTRIBUTIONS [J].
ALCALA, JR ;
GRATTON, E ;
PRENDERGAST, FG .
BIOPHYSICAL JOURNAL, 1987, 51 (06) :925-936
[2]  
Andrade SM, 2002, BIOPHYS J, V82, P1607, DOI 10.1016/S0006-3495(02)75512-4
[3]  
Andrade SM, 2000, PHOTOCHEM PHOTOBIOL, V72, P444, DOI 10.1562/0031-8655(2000)072<0444:TLOTNA>2.0.CO
[4]  
2
[5]   Kinetic refolding of β-lactoglobulin.: Studies by synchrotron X-ray scattering, and circular dichroism, absorption and fluorescence spectroscopy [J].
Arai, M ;
Ikura, T ;
Semisotnov, GV ;
Kihara, H ;
Amemiya, Y ;
Kuwajima, K .
JOURNAL OF MOLECULAR BIOLOGY, 1998, 275 (01) :149-162
[6]   Effect of pH on the structure and aggregation of human glycodelin A.: A comparison with β-lactoglobulin A [J].
Barteri, M ;
Gaudiano, MC ;
Rotella, S ;
Benagiano, G ;
Pala, A .
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEIN STRUCTURE AND MOLECULAR ENZYMOLOGY, 2000, 1479 (1-2) :255-264
[7]   POLARITY OF AOT MICELLAR INTERFACES - USE OF THE PREFERENTIAL SOLVATION CONCEPTS IN THE EVALUATION OF THE EFFECTIVE DIELECTRIC-CONSTANTS [J].
BELLETETE, M ;
LACHAPELLE, M ;
DUROCHER, G .
JOURNAL OF PHYSICAL CHEMISTRY, 1990, 94 (13) :5337-5341
[8]   EXCIPLEX FORMATION BETWEEN PYRENE DERIVATIVES AND N,N-DIMETHYLANILINE IN AEROSOL-OT REVERSED MICELLES [J].
BORSARELLI, CD ;
COSA, JJ ;
PREVITALI, CM .
LANGMUIR, 1992, 8 (04) :1070-1075
[9]   Effect of water-soluble polymers on the morphology of aerosol OT vesicles [J].
Briz, JI ;
Velázquez, MM .
JOURNAL OF COLLOID AND INTERFACE SCIENCE, 2002, 247 (02) :437-446
[10]   TIME RESOLVED SPECTROSCOPY OF TRYPTOPHYL FLUORESCENCE OF ESCHERICHIA-COLI LAC REPRESSOR [J].
BROCHON, JC ;
WAHL, P ;
CHARLIER, M ;
MAURIZOT, JC ;
HELENE, C .
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 1977, 79 (04) :1261-1271