光谱法和分子对接模拟技术研究异烟肼对人血清白蛋白和过氧化氢酶的特异性结合及抑制作用

异烟肼(Isoniazid, INH)是最常用的一线抗结核药物之一。据报道，人体内高浓度的异烟肼可导致癫痫, 肝功能衰竭, 甚至死亡。因此，研究异烟肼对人血清白蛋白(HSA)和过氧化氢酶(CAT)的结构和活性的潜在结合影响有利于评估其毒性和副作用。在模拟生理条件下，利用多种荧光光谱和分子对接技术研究INH与HSA和CAT之间的相互作用。所有荧光数据均进行了内滤光校正以获得更准确的结合参数。结果表明，INH-HSA和INH-CAT体系的猝灭常数(K_sv)随着温度的升高而降低，表明INH对HSA及CAT的荧光猝灭机理为静态猝灭。利用紫外-可见吸收光谱、同步荧光光谱、和圆二色(CD)光谱法研究了INH对HSA和CAT构象的影响。结果发现，INH可改变色氨酸残基的微环境并降低HSA和CAT中α-螺旋结构，导致蛋白质结构发生伸展，进而可能影响其生理功能。分子对接结果表明，INH与HSA的结合位点位于HSA的site Ⅰ，ⅡA子域。INH可以进入CAT中β-折叠的桶状空腔，从而抑制CAT的活性。Hill系数结果表明，INH与左氧氟沙星(LVFX，一种安全有效的二线抗结核药物，与其他抗结核药物联合使用可以提高抗结核疗效)之间存在药物协同性，促进INH与HSA的相互作用。另外，CD光谱测定表明INH与LVFX的协同作用改变HSA的二级结构，使α-螺旋结构降低约7.9%。该研究探讨了INH与HSA和CAT之间的结合作用和毒性机制，为INH的安全使用提供重要依据。

Spectroscopic and Molecular Docking Study on Specific Binding and Inhibition of Isoniazid to Human Serum Albumin and Catalase

Isonicotinic acid hydrazide (Isoniazid,INH)is one of the most commonly used first-line anti-tuber-culosis drugs,which has been reported that the high concentration of INH in human body can lead to epilepsy, liver function failure,and even death.Therefore,studying the potential binding effects of INH on the struc-ture and activity of human serum albumin (HSA)and catalase (CAT)is very essential for evaluating its toxici-ty and side effect.In this paper,multi-spectroscopic and molecular docking methods were used to elucidate the patterns of INH to HSA and CAT under imitated physiological conditions.The inner filter effect of all fluores-cence data in the paper was eliminated to get accurate binding parameters.The Stern-Volmer quenching con-stants (KSV)of both HSA-INH system and CAT-INH system inversely correlated with temperatures,demon-strating that INH quench the intrinsic fluorescence of HSA and CAT via static quenching.The conformational investigation of HSA and CAT through UV-visible absorption spectroscopy,synchronous fluorescence and cir-cular dichroism (CD)showed that INH could change the micro-environment of tryptophan residues and re-duced theα-helix content of protein.These results demonstrated that the binding of INH may lead to the loos-ening of protein skeleton,which which may affect its physiological function.The results of molecular docking revealed that the INH was located in Sudlow’s site I of HSA.And INH bound to CAT at a cavity among the wrapping domain helical domain andβ-barrel,which resulted in the inhibition of CAT activity.In addition, Levofloxacin (LVFX)is a new effective and safe second-line anti-tuberculosis drugs and can improve the cura-tive effect on anti-TB by using with other anti-TB drugs,the result of Hill’s coefficients (nH )about synergy between INH and proved that LVFX promoted the interaction of HSA with INH.Moreover,according to the CD spectra,synergy between INH and LVFX changed the conformation of HSA and the amount ofα-helix de-creased about 7.9%.This work will provide important insights into the binding and toxicity mechanism of INH to HSA and CAT in vivo and is expected to be helpful in evaluating the essential information for using the INH safely.