Effect of Tb3+ on the Stability of Ovalbumin†

doi:10.7503/cjcu20160259

Abstract

Abstract:

The binding properties of Tb³⁺ with ovalbumin were measured by fluorescence spectra at room temperature in pH=7.4, 10 mmol/L 4-(2-hydroxyethyl)-1-piperazinecthanesulfonic acid(Hepes). The effect of Tb³⁺ on the hydrophobic region of ovalbumin was measured by hydrophobic probe 2-p-toluidino-6-naphthalenesalfonic acid(TNS). The effects of Ca²⁺ and Tb³⁺ on the stability of ovalbumin were analyzed by chemical unfolding methods. The results suggested that ovalbumin can form 1∶2 complexes with Tb³⁺. The binding of Tb³⁺ increased the hydrophobic surface of ovalbumin, and the fluorescence intensity of TNS increased. The unfolding curves of ovalbumin showed two transition, N $?$ I and I $?$ U. Ca²⁺ can increase the stability of transition I $?$ U and Tb³⁺ can increase the stability of transitions N $?$ I and I $?$ U.

Key words: Ovalbumin, Tb3+, Stability(Ed.: F, K, M)

CLC Number:

O614
O629.73

TrendMD:

SONG Zhen, DONG Jinlong, REN Yuehong, YUAN Wen, ZHANG Caifeng, YANG Binsheng. Effect of Tb³⁺ on the Stability of Ovalbumin^†[J]. Chem. J. Chinese Universities, 2016, 37(7): 1245.

Figures/Tables 7

Fig.1 Structure of Ova(PDB 1Ova) drawn by using Pymol software Ca2+ was shown as sphere, Trp was shown as sticks.

Fig.2 Fluorescence spectra of Ova at different concentrations of Tb3+ in pH=7.4, 10 mmol/L

Fig.3 Fluorescence spectra of Ova at different concentrations of TNS in the absence of Ova, in pH=7.4, 10 mmol/L Hepes(A), titration curve of Ova with TNS addition(B) and fluorescence spectra of TNS with the addition of Tb3+ in the presence of Ova in pH=7.4, 10 mmol/L Hepes(C)

Fig.4 Fluorescence spectra for protein in different concentrations of urea(A) and the unfolding curves of

Table 1 Thermodynamics parameters of proteins from urea-induced unfolding at pH=7.4 and 25 ℃*

Protein	N I		I U					<Δ $G element 0$ >/ (kJ·mol^-1)
Protein	[D]_1/2/ (mol·L^-1)	m/ (kJ·mol^-2·L)	Δ $G i 0$ / (kJ·mol^-1)		[D]_1/2/ (mol·L^-1)	m/ (kJ·mol^-2·L)	Δ $G i 0$ / (kJ·mol^-1)
Ova	4.02	-3.38±0.39	13.62±0.26	7.16	-2.63±0.19	18.83±0.11		16.35±0.13
Ova-Ca²⁺	4.07	-3.33±0.15	13.57±0.76	8.03	-2.63±0.06	21.18±0.24		18.13±0.27
Ova-Tb³⁺	5.16	-3.05±0.38	15.74±0.26	8.22	-2.68±0.13	22.08±0.26		19.54±0.21

Table 1 Thermodynamics parameters of proteins from urea-induced unfolding at pH=7.4 and 25 ℃*

Protein	N I		I U					<Δ $G element 0$ >/ (kJ·mol^-1)
Protein	[D]_1/2/ (mol·L^-1)	m/ (kJ·mol^-2·L)	Δ $G i 0$ / (kJ·mol^-1)		[D]_1/2/ (mol·L^-1)	m/ (kJ·mol^-2·L)	Δ $G i 0$ / (kJ·mol^-1)
Ova	4.02	-3.38±0.39	13.62±0.26	7.16	-2.63±0.19	18.83±0.11		16.35±0.13
Ova-Ca²⁺	4.07	-3.33±0.15	13.57±0.76	8.03	-2.63±0.06	21.18±0.24		18.13±0.27
Ova-Tb³⁺	5.16	-3.05±0.38	15.74±0.26	8.22	-2.68±0.13	22.08±0.26		19.54±0.21

Fig.5 Temperature induced unfolding of Ova, Ova-Ca2+, Ova-Tb3+ in pH=7.4, 10 mmol/L Hepes

Table 2 Thermal denaturation parameters of proteins at pH=7.4 and 25 ℃*

Protein	N I		I U		ΔH_m/(kJ·mol^-1)
Protein	T_m/℃	Δ $H m 1$ /(kJ·mol^-1)		T_m/℃	Δ $H m 2$ /(kJ·mol^-1)
Ova	42.34	5.23±0.14	70.69	9.74±0.41	7.64
Ova-Ca²⁺	42.89	5.45±0.12	75.67	12.13±0.09	9.46
Ova-Tb³⁺	48.75	7.59±0.21	79.18	13.29±0.27	11.01

Table 2 Thermal denaturation parameters of proteins at pH=7.4 and 25 ℃*

Protein	N I		I U		ΔH_m/(kJ·mol^-1)
Protein	T_m/℃	Δ $H m 1$ /(kJ·mol^-1)		T_m/℃	Δ $H m 2$ /(kJ·mol^-1)
Ova	42.34	5.23±0.14	70.69	9.74±0.41	7.64
Ova-Ca²⁺	42.89	5.45±0.12	75.67	12.13±0.09	9.46
Ova-Tb³⁺	48.75	7.59±0.21	79.18	13.29±0.27	11.01

References 14

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Effect of Tb³⁺ on the Stability of Ovalbumin^†

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