Meclofenoxate Inhibits Aggregation of Alpha-synuclein in vitro.

IF 1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY
Adhuna Parui, Soumojit Biswas, Ipsita Roy
{"title":"Meclofenoxate Inhibits Aggregation of Alpha-synuclein <i>in vitro</i>.","authors":"Adhuna Parui,&nbsp;Soumojit Biswas,&nbsp;Ipsita Roy","doi":"10.2174/0929866530666230307113055","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>α-Synuclein, a natively disordered protein, is a key component of Lewy bodies, the ubiquitinated protein aggregates which are the pathological hallmark of Parkinson's disease (PD). Meclofenoxate (centrophenoxine) is a nootropic drug which has shown beneficial therapeutic effects in various neuronal diseases. Administration of meclofenoxate enhanced levels of dopamine and improved motor function in animal models of Parkinson's disease (PD). Evidence suggested that dopamine interacts with and modulates α-synuclein aggregation.</p><p><strong>Objective: </strong>The aim of this work was to investigate whether the observed positive effect of addition of meclofenoxate, a nootropic agent, on dopamine level, could be correlated with its effect on aggregation of α-synuclein.</p><p><strong>Methods: </strong>Purification of recombinant human α-synuclein was performed by anion exchange chromatography. The purified protein was incubated in the absence and presence of meclofenoxate and was analyzed for aggregation by Thioflavin T fluorescence spectroscopy. Conformational changes in α-synuclein were monitored by fluorescence spectroscopy and fluorescence quenching studies using a neutral quencher. Secondary structure analysis of α-synuclein was monitored by circular dichroism spectroscopy.</p><p><strong>Results: </strong>Recombinant human α-synuclein was expressed and purified by anion-exchange chromatography. Incubation of α-synuclein with meclofenoxate led to lowering aggregation in a concentration-dependent manner. Reduction in formation of oligomers was seen which suggested the formation of an off-pathway species which did not give rise to an aggregation-competent entity. Fluorescence quenching studies revealed that the additive distorted the native conformation of α- synuclein, leading to the formation of lower amounts of aggregation-prone species.</p><p><strong>Conclusion: </strong>In the presence of higher concentrations of meclofenoxate, α-synuclein undergoes a change in its conformation. This change is not dependent on the concentration of the additive. This non-native conformer promotes the formation of a species which does not undergo further aggregation. Our study provides a mechanistic explanation of the earlier observation that meclofenoxate has a beneficial effect on progression of PD in animal models.</p>","PeriodicalId":20736,"journal":{"name":"Protein and Peptide Letters","volume":"30 5","pages":"361-366"},"PeriodicalIF":1.0000,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Protein and Peptide Letters","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.2174/0929866530666230307113055","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0

Abstract

Background: α-Synuclein, a natively disordered protein, is a key component of Lewy bodies, the ubiquitinated protein aggregates which are the pathological hallmark of Parkinson's disease (PD). Meclofenoxate (centrophenoxine) is a nootropic drug which has shown beneficial therapeutic effects in various neuronal diseases. Administration of meclofenoxate enhanced levels of dopamine and improved motor function in animal models of Parkinson's disease (PD). Evidence suggested that dopamine interacts with and modulates α-synuclein aggregation.

Objective: The aim of this work was to investigate whether the observed positive effect of addition of meclofenoxate, a nootropic agent, on dopamine level, could be correlated with its effect on aggregation of α-synuclein.

Methods: Purification of recombinant human α-synuclein was performed by anion exchange chromatography. The purified protein was incubated in the absence and presence of meclofenoxate and was analyzed for aggregation by Thioflavin T fluorescence spectroscopy. Conformational changes in α-synuclein were monitored by fluorescence spectroscopy and fluorescence quenching studies using a neutral quencher. Secondary structure analysis of α-synuclein was monitored by circular dichroism spectroscopy.

Results: Recombinant human α-synuclein was expressed and purified by anion-exchange chromatography. Incubation of α-synuclein with meclofenoxate led to lowering aggregation in a concentration-dependent manner. Reduction in formation of oligomers was seen which suggested the formation of an off-pathway species which did not give rise to an aggregation-competent entity. Fluorescence quenching studies revealed that the additive distorted the native conformation of α- synuclein, leading to the formation of lower amounts of aggregation-prone species.

Conclusion: In the presence of higher concentrations of meclofenoxate, α-synuclein undergoes a change in its conformation. This change is not dependent on the concentration of the additive. This non-native conformer promotes the formation of a species which does not undergo further aggregation. Our study provides a mechanistic explanation of the earlier observation that meclofenoxate has a beneficial effect on progression of PD in animal models.

甲氯芬诺酯体外抑制α -突触核蛋白聚集。
背景:α-突触核蛋白是一种天然紊乱蛋白,是路易小体的关键组成部分,路易小体是泛素化蛋白聚集体,是帕金森病(PD)的病理标志。甲氯芬诺酯(正苯诺辛)是一种益智药物,在多种神经疾病中显示出良好的治疗效果。在帕金森病(PD)动物模型中,给予甲氯芬诺酯可提高多巴胺水平并改善运动功能。有证据表明,多巴胺与α-突触核蛋白聚集相互作用并调节其聚集。目的:探讨促智剂甲氯芬诺酯对多巴胺水平的积极影响是否与其对α-突触核蛋白聚集的影响有关。方法:采用阴离子交换色谱法纯化重组人α-突触核蛋白。纯化后的蛋白在不存在和不存在甲氯虫腈的情况下孵育,用硫黄素T荧光光谱分析其聚集性。利用荧光光谱和中性猝灭剂的荧光猝灭研究监测α-突触核蛋白的构象变化。采用圆二色光谱法对α-synuclein的二级结构进行分析。结果:表达了重组人α-突触核蛋白,并通过阴离子交换层析纯化。α-突触核蛋白与甲氯膦酸酯孵育导致以浓度依赖的方式降低聚集。低聚物的形成减少,这表明形成了一个通路外的物种,没有产生聚集能力的实体。荧光猝灭研究表明,添加剂扭曲了α-突触核蛋白的天然构象,导致形成较低数量的易聚集物质。结论:在较高浓度的甲氯虫腈存在下,α-突触核蛋白的构象发生改变。这种变化不依赖于添加剂的浓度。这种非本地的构象促进了不进行进一步聚集的物种的形成。我们的研究为早期观察提供了一种机制解释,即甲氯膦酸盐对动物模型中PD的进展有有益的影响。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
Protein and Peptide Letters
Protein and Peptide Letters 生物-生化与分子生物学
CiteScore
2.90
自引率
0.00%
发文量
98
审稿时长
2 months
期刊介绍: Protein & Peptide Letters publishes letters, original research papers, mini-reviews and guest edited issues in all important aspects of protein and peptide research, including structural studies, advances in recombinant expression, function, synthesis, enzymology, immunology, molecular modeling, and drug design. Manuscripts must have a significant element of novelty, timeliness and urgency that merit rapid publication. Reports of crystallization and preliminary structure determination of biologically important proteins are considered only if they include significant new approaches or deal with proteins of immediate importance, and preliminary structure determinations of biologically important proteins. Purely theoretical/review papers should provide new insight into the principles of protein/peptide structure and function. Manuscripts describing computational work should include some experimental data to provide confirmation of the results of calculations. Protein & Peptide Letters focuses on: Structure Studies Advances in Recombinant Expression Drug Design Chemical Synthesis Function Pharmacology Enzymology Conformational Analysis Immunology Biotechnology Protein Engineering Protein Folding Sequencing Molecular Recognition Purification and Analysis
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信