Drug-receptor interaction plays an important role in a series of biological effects, such as cell pro- liferation, immune response, tumor metastasis, and drug delivery. Therefore, the research on drug-re- ceptor inter...Drug-receptor interaction plays an important role in a series of biological effects, such as cell pro- liferation, immune response, tumor metastasis, and drug delivery. Therefore, the research on drug-re- ceptor interaction is growing rapidly. The equilibrium dissociation constant （KD） is the basic parameter to evaluate the binding property of the drug-receptor. Thus, a variety of analytical methods have been established to determine the KD values, including radioligand binding assay, surface plasmon resonance method, fluorescence energy resonance transfer method, affinity chromatography, and isothermal ti- tration calorimetry. With the invention and innovation of new technology and analysis method, there is a deep exploration and comprehension about drug-receptor interaction. This review discusses the differ- ent methods of determining the KD values, and analyzes the applicability and the characteristic of each analytical method. Conclusively, the aim is to provide the guidance for researchers to utilize the most appropriate analytical tool to determine the KD values.展开更多
Equilibrium constants are essential for understanding and predicting the behavior of chemical systems across various scientifc disciplines.Traditionally,these constants are computed via nonlinear regression of reactio...Equilibrium constants are essential for understanding and predicting the behavior of chemical systems across various scientifc disciplines.Traditionally,these constants are computed via nonlinear regression of reaction isotherms,which show the depend-ence of the unreacted fraction of one reactant on the total concentration of another reactant.However,while these equilibrium constants can be precise(with small random errors),they may also be grossly inaccurate(with large systematic errors),leading to potential misinterpretations.Although some statisticalmethods exist for assessing the accuracy of nonlinear regression,their limitedpracticality for molecular scientists has resulted in their neglect by this research community.The objective of this work is to develop a practical method for quantitatively assessing the accuracy of equilibrium constants that could be easily understood and immediately adopted by researchers routinely determining these constants.Our approach integrates error-propagation and regression-stability analyses to establish the accuracy confidence interval(ACI)-a range within which the true value of the computed parameter lies with a defined probability.In a proof-of-principle study,we applied this approach to develop a workflow for determining the ACI of the equilibrium dissociation constant(K_(d))of affinity complexes from a single binding isotherm.We clearly explained how the input parameters for this workflow can be determined,and finally,we have implemented this workflow in a user-friendly web application(https://aci.sci.yorku.ca)to facilitate its immediate adoption by molecular scientists,regardless of their mathematical and computer proficiency.We further conducted three case studies exemplifying the use of the ACI in the context of simultaneous assessment of precision and accuracy of determined K_(d)values.By understanding the ACI of equilibrium constants and other parameters computed through nonlinear regression,researchers can avoid misconceptions that arise from relying solely on precision.展开更多
Upon the study of small-molecules binding to proteins, the traditional methods for calculating dissociation constants (Kd and Ki) have shortcomings in dealing with the single bind- ing site models. In this paper, two ...Upon the study of small-molecules binding to proteins, the traditional methods for calculating dissociation constants (Kd and Ki) have shortcomings in dealing with the single bind- ing site models. In this paper, two equations have been derived to solve this problem. These two equations are independent of the total concentration or initial degree of saturation of receptor and the activity of the competitive molecule. Through nonlinear fitting against these two equations, Kd value of a probe can be obtained by binding assay, and Ki value of a ligand can be obtained by competitive assay. Moreover, only the total concentrations of receptor([R]t), ligand([L]t) and probe([P]t) are required for the data fitting. In this work, Ki values of some typical ligands of PPARγ were successfully determined by use of our equations, among which the Ki value of PPARγ-LY171883 was reported for the first time.展开更多
Twenty four male guinea-pigs were randomly divided into 3 groups: Group A, allergic asthmaguinea-pigs prepared by peritoneal injection and spraying inhalation of albumin; Group B, allergic asthmaguinea-pigs treated wi...Twenty four male guinea-pigs were randomly divided into 3 groups: Group A, allergic asthmaguinea-pigs prepared by peritoneal injection and spraying inhalation of albumin; Group B, allergic asthmaguinea-pigs treated with electroacupuncture through stimulating the acupoint Feishu (UB13 ) and Group C, thecontrol group. With[3H] -DHA as a radioligand, pulmonary adrenoceptor ( AC) in the 3 groups of guinea-pigs were detrermined by radioligand binding assay technique. The results showed that: ( 1 ) The maximumbinding volume of receptor ligand ( Bmax, fmol/mg prot. ) of pulmonary AC in Group A (123. 86 42. 91 )was significantly lower than that in group C (199 . 54 37. 03 , P< 0. 05) , while the difference between groupB ( 142. 00 42. 91 ) and group C was not significant ( P > 0. 05 ) . ( 2 ) The Kd ( nM) of pulmonary AC ingroup A, B and C were 5 . 10 1 . 39 , 6. 53 2 . 41 and 8. 72 2 . 02 respectively, and the differences amongthe three groups were not significant ( P >0. 05) . The results indicated that the decrease of the content anddysfunction of pulmonary AC in allergic asthma guinea-pigs might be regulated by electroacupuncture thera-py.展开更多
文摘Drug-receptor interaction plays an important role in a series of biological effects, such as cell pro- liferation, immune response, tumor metastasis, and drug delivery. Therefore, the research on drug-re- ceptor interaction is growing rapidly. The equilibrium dissociation constant （KD） is the basic parameter to evaluate the binding property of the drug-receptor. Thus, a variety of analytical methods have been established to determine the KD values, including radioligand binding assay, surface plasmon resonance method, fluorescence energy resonance transfer method, affinity chromatography, and isothermal ti- tration calorimetry. With the invention and innovation of new technology and analysis method, there is a deep exploration and comprehension about drug-receptor interaction. This review discusses the differ- ent methods of determining the KD values, and analyzes the applicability and the characteristic of each analytical method. Conclusively, the aim is to provide the guidance for researchers to utilize the most appropriate analytical tool to determine the KD values.
基金supported by a York University grant to the Catalyzing Interdisciplinary Research Cluster“Technologies for Identification and Control of Infectious Diseases”and an NSERC Discovery Grant(RGPIN-2022-04563)to S.N.KAdditional support was provided by an NSERC Undergraduate Student Research Award(USRA-582491-2023)to J.La University of Münster visiting fellowship to S.N.K.We extend our gratitude to Prof.Howard A.Levin and Dr.Yeon-Jung Seo from Iowa State University and Prof.Jianhong Wu from York University for their valuable assistance in refining the manuscript.
文摘Equilibrium constants are essential for understanding and predicting the behavior of chemical systems across various scientifc disciplines.Traditionally,these constants are computed via nonlinear regression of reaction isotherms,which show the depend-ence of the unreacted fraction of one reactant on the total concentration of another reactant.However,while these equilibrium constants can be precise(with small random errors),they may also be grossly inaccurate(with large systematic errors),leading to potential misinterpretations.Although some statisticalmethods exist for assessing the accuracy of nonlinear regression,their limitedpracticality for molecular scientists has resulted in their neglect by this research community.The objective of this work is to develop a practical method for quantitatively assessing the accuracy of equilibrium constants that could be easily understood and immediately adopted by researchers routinely determining these constants.Our approach integrates error-propagation and regression-stability analyses to establish the accuracy confidence interval(ACI)-a range within which the true value of the computed parameter lies with a defined probability.In a proof-of-principle study,we applied this approach to develop a workflow for determining the ACI of the equilibrium dissociation constant(K_(d))of affinity complexes from a single binding isotherm.We clearly explained how the input parameters for this workflow can be determined,and finally,we have implemented this workflow in a user-friendly web application(https://aci.sci.yorku.ca)to facilitate its immediate adoption by molecular scientists,regardless of their mathematical and computer proficiency.We further conducted three case studies exemplifying the use of the ACI in the context of simultaneous assessment of precision and accuracy of determined K_(d)values.By understanding the ACI of equilibrium constants and other parameters computed through nonlinear regression,researchers can avoid misconceptions that arise from relying solely on precision.
基金the National Natural Science Foundation of China(Grants No.20372069)the 863 Hi-Tech Program(Grant Nos.2002AA233011,2003AA235030,2001AA235071).
文摘Upon the study of small-molecules binding to proteins, the traditional methods for calculating dissociation constants (Kd and Ki) have shortcomings in dealing with the single bind- ing site models. In this paper, two equations have been derived to solve this problem. These two equations are independent of the total concentration or initial degree of saturation of receptor and the activity of the competitive molecule. Through nonlinear fitting against these two equations, Kd value of a probe can be obtained by binding assay, and Ki value of a ligand can be obtained by competitive assay. Moreover, only the total concentrations of receptor([R]t), ligand([L]t) and probe([P]t) are required for the data fitting. In this work, Ki values of some typical ligands of PPARγ were successfully determined by use of our equations, among which the Ki value of PPARγ-LY171883 was reported for the first time.
文摘Twenty four male guinea-pigs were randomly divided into 3 groups: Group A, allergic asthmaguinea-pigs prepared by peritoneal injection and spraying inhalation of albumin; Group B, allergic asthmaguinea-pigs treated with electroacupuncture through stimulating the acupoint Feishu (UB13 ) and Group C, thecontrol group. With[3H] -DHA as a radioligand, pulmonary adrenoceptor ( AC) in the 3 groups of guinea-pigs were detrermined by radioligand binding assay technique. The results showed that: ( 1 ) The maximumbinding volume of receptor ligand ( Bmax, fmol/mg prot. ) of pulmonary AC in Group A (123. 86 42. 91 )was significantly lower than that in group C (199 . 54 37. 03 , P< 0. 05) , while the difference between groupB ( 142. 00 42. 91 ) and group C was not significant ( P > 0. 05 ) . ( 2 ) The Kd ( nM) of pulmonary AC ingroup A, B and C were 5 . 10 1 . 39 , 6. 53 2 . 41 and 8. 72 2 . 02 respectively, and the differences amongthe three groups were not significant ( P >0. 05) . The results indicated that the decrease of the content anddysfunction of pulmonary AC in allergic asthma guinea-pigs might be regulated by electroacupuncture thera-py.
