Comparisons and Applications of Functional Equations for the Calculation of the Protein-Ligand Binding Constant Based on Fluorescence Spectral Data

Jing ZHANG; Xuan GAO; Liang JIN; Hong-hui WANG; Xi-ping ZHOU

doi:10.3964/j.issn.1000-0593(2020)11-3494-05

Journals >Spectroscopy and Spectral Analysis >Volume 40 >Issue 11 >Page 3494 > Article

Spectroscopy and Spectral Analysis
Vol. 40, Issue 11, 3494 (2020)

Comparisons and Applications of Functional Equations for the Calculation of the Protein-Ligand Binding Constant Based on Fluorescence Spectral Data

Jing ZHANG^1、1, Xuan GAO^1、1, Liang JIN^1、1, Hong-hui WANG^1、1, and Xi-ping ZHOU^1、1

Author Affiliations

¹[in Chinese]

¹1. Key Laboratory of Estuarine Ecological Security and Environmental Health, School of Environmental Science and Engineering, Xiamen University Tan Kah Kee College, Zhangzhou 363105, China

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DOI: 10.3964/j.issn.1000-0593(2020)11-3494-05 Cite this Article

Jing ZHANG, Xuan GAO, Liang JIN, Hong-hui WANG, Xi-ping ZHOU. Comparisons and Applications of Functional Equations for the Calculation of the Protein-Ligand Binding Constant Based on Fluorescence Spectral Data[J]. Spectroscopy and Spectral Analysis, 2020, 40(11): 3494 Copy Citation Text

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(a) Job’s plot for fluorescence quenching of HSA-NFX system at 282 nm excitation with total concentration of 2.0×10-5 mol·L-1 ([HSA]+[NFX]); (b) The relative fluorescence intensity of HSA in the presence of various concentrations of NFXEx=282.0 nm, Em=332.0 nm

Fig. 1. (a) Job’s plot for fluorescence quenching of HSA-NFX system at 282 nm excitation with total concentration of 2.0×10^-5 mol·L^-1 ([HSA]+[NFX]); (b) The relative fluorescence intensity of HSA in the presence of various concentrations of NFX
Ex=282.0 nm, Em=332.0 nm

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结合比	假设a	假设b	The optimal equation^*
1:1	成立	成立	$\frac{F_{0} - F}{F}$ =K_b[L_add]	(S6)
	不成立	成立	$\frac{F_{0} - F}{F - F_{c}}$ =K_b[L_add]	(S7)
			$\frac{F_{0}}{F_{0} - F}$ = $\frac{1}{f_{a} K_{b} [L_{add}]}$ + $\frac{1}{f_{a}}$	(S8)
	成立	不成立	$\frac{F_{0} - F}{F_{0}}$ = $\frac{[P_{add}] + [L_{add}] + \frac{1}{K_{b}} - \sqrt[]{{([P_{add}] + [L_{add}] + \frac{1}{K_{b}})}^{2} - 4 [P_{add}] [L_{add}]}}{2 [P_{add}]}$	(S12)
	不成立	不成立	$\frac{F_{0} - F}{F_{0} - F_{c}}$ = $\frac{[P_{add}] + [L_{add}] + \frac{1}{K_{b}} - \sqrt[]{{([P_{add}] + [L_{add}] + \frac{1}{K_{b}})}^{2} - 4 [P_{add}] [L_{add}]}}{2 [P_{add}]}$	(S13)
1:n (n≥2)	成立	成立	lg $\frac{F_{0} - F}{F}$ =lgK_b+nlg[L_add]	(S17)
	不成立	成立	lg $\frac{F_{0} - F}{F - F_{c}}$ =lgK_b+nlg[L_add]	(S18)
	成立	不成立	lg $\frac{F_{0} - F}{F}$ =lgK_b+nlg $([L_{add}] - n [P_{add}] \frac{F_{0} - F}{F_{0}})$	(S19)
			$\frac{F_{0}}{F}$ = $\frac{K_{b} [L_{add}] F_{0}}{F_{0} - F}$ -nK_b[P_add]	(S22)
	不成立	不成立	lg $\frac{F_{0} - F}{F - F_{c}}$ =lgK_b+nlg $([L_{add}] - n [P_{add}] \frac{F_{0} - F}{F_{0} - F_{c}})$	(S20)
			lg $\frac{F_{0} - F}{F - F_{c}}$ =nlgK_a+nlg $([L_{add}] - n [P_{add}] \frac{F_{0} - F}{F_{0} - F_{c}})$	(S21)
			$\frac{F_{0} - F_{c}}{F - F_{c}}$ = $\frac{K_{b} [L_{add}] (F_{0} - F_{c})}{F_{0} - F}$ -nK_b[P_add]	(S23)
			K_b= $\frac{β}{1 - β}$ × $\frac{1}{[L]}$	(S24)

Table 1. Selection of equations for 1:1 or 1:n protein-ligand binding processes

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分析方法	温度/K	K_b/(L·mol^-1)	参考文献
荧光光谱法	299.15	5.97×10⁴	[8]
	298	2.21×10⁷	[5]
	298	4.68×10⁴	[9]
	298	5.0×10⁴	This work
表面等离子体共振法	298.15	(1.00~1.04)×10³	[10]
等温滴定量热法	298	(3.49~3.85)×10⁴	[11]
亲和毛细管电泳法	298	6.35×10⁴	[9]

Table 2. The binding constant of HSA-NFX system obtained by different analytical techniques

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