红麻种子半胱氨酸蛋白酶(KSCP)抑制凝血级联和血小板聚集的内在途径。

IF 16.4 1区 化学 Q1 CHEMISTRY, MULTIDISCIPLINARY
Sujatha M Hanumegowda, Chandramma Srinivasa, Ashwini Shivaiah, Manjula M Venkatappa, Rohith L Shankar, Ramesh K Lakshmaiah, Sathisha J Gonchigar, Devaraja Sannaningaiah
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引用次数: 0

摘要

背景:血栓形成是阻碍血液在整个循环系统中流动的关键事件,可导致中风、心肌梗死和严重的心血管并发症。目前,可用的抗血栓药物会引发几种危及生命的副作用。导论:天然来源的抗血栓药物无不良反应,引起了人们的高度关注。在我们之前的研究中,我们报道了红麻种子蛋白提取物的抗氧化、抗凝血和抗血小板性能。因此,在目前的研究中,从红麻种子蛋白提取物中纯化和表征具有潜在抗血栓活性的半胱氨酸蛋白酶。方法:采用凝胶渗透法和离子交换柱层析法纯化红麻种子半胱氨酸蛋白酶。用十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS PAGE)对酶的纯度进行鉴定。采用反相高效液相色谱法(RP-HPLC)、基质辅助激光解吸电离飞行时间法(MALDI-TOF)和圆二色技术(CD)对其进行表征。利用光谱学对其进行了蛋白水解、纤维蛋白溶解和动力学研究。采用血浆再钙化时间、凝血酶原时间(PT)、凝血酶凝血时间(TCT)、活化部分凝血活素时间(APTT)、出血时间和血小板聚集研究KSCP的抗血栓活性。结果:在还原和非还原条件下均可观察到KSCP单条尖锐带,分子量为24.1667kDa。发现KSCP含有30.3%的螺旋旋转和69.7%的随机线圈,没有β褶板。KSCP消化酪蛋白和纤维蛋白,其活性受到碘乙酸(IAA)的抑制。KSCP在pH 6.0、温度40℃时活性最佳。KSCP通过干扰血凝级联的内在途径表现出抗凝特性。此外,KSCP溶解全血和血浆凝块和血小板聚集。结论:从红麻籽提取物中纯化的KSCP具有抗血栓作用。因此,它可能是一个更好的候选管理血栓性并发症。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Kenaf Seed Cysteine Protease (KSCP) Inhibits the Intrinsic Pathway of the Blood Coagulation Cascade and Platelet Aggregation.

Background: Thrombosis is the key event that obstructs the flow of blood throughout the circulatory system, leading to stroke, myocardial infarction and severe cardiovascular complications. Currently, available antithrombotic drugs trigger several life-threatening side effects.

Introduction: Antithrombotic agents from natural sources devoid of adverse effects are grabbing high attention. In our previous study, we reported the antioxidant, anticoagulant and antiplatelet properties of kenaf seed protein extract. Therefore, in the current study, purification and characterization of cysteine protease from kenaf seed protein extract responsible for potential antithrombotic activity was undertaken.

Methods: Purification of KSCP (Kenaf Seed Cysteine Protease) was carried out using gel permeation and ion exchange column chromatography. The purity of the enzyme was evaluated by SDS PAGE (Sodium Dodecyl-Sulfate Polyacrylamide Gel Electrophoresis). RP-HPLC (Reverse Phase High-Performance Liquid Chromatography), MALDI-TOF (Matrix-Assisted Laser Desorption Ionization Time-Of-Flight) and CD (Circular Dichroism techniques) were employed for its characterization. Proteolytic, fibrinolytic and kinetic study was done using spectroscopy. Plasma recalcification time, Prothrombin Time (PT), Thrombin clotting time (TCT), Activated Partial Thromboplastin Time (APTT), bleeding time and platelet aggregation studies were carried out for antithrombotic activity of KSCP.

Result: A single sharp band of KSCP was observed under both reduced and non-reduced conditions, having a molecular mass of 24.1667kDa. KSCP was found to contain 30.3% helix turns and 69.7% random coils without a beta-pleated sheet. KSCP digested casein and fibrin, and its activity was inhibited by iodoacetic acid (IAA). KSCP was optimally active at pH 6.0 at the temperature of 40°C. KSCP exhibited anticoagulant properties by interfering in the intrinsic pathway of the blood coagulation cascade. Furthermore, KSCP dissolved both whole blood and plasma clots and platelet aggregation.

Conclusion: KSCP purified from kenaf seed extract showed antithrombotic potential. Hence, it could be a better candidate for the management of thrombotic complications.

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来源期刊
Accounts of Chemical Research
Accounts of Chemical Research 化学-化学综合
CiteScore
31.40
自引率
1.10%
发文量
312
审稿时长
2 months
期刊介绍: Accounts of Chemical Research presents short, concise and critical articles offering easy-to-read overviews of basic research and applications in all areas of chemistry and biochemistry. These short reviews focus on research from the author’s own laboratory and are designed to teach the reader about a research project. In addition, Accounts of Chemical Research publishes commentaries that give an informed opinion on a current research problem. Special Issues online are devoted to a single topic of unusual activity and significance. Accounts of Chemical Research replaces the traditional article abstract with an article "Conspectus." These entries synopsize the research affording the reader a closer look at the content and significance of an article. Through this provision of a more detailed description of the article contents, the Conspectus enhances the article's discoverability by search engines and the exposure for the research.
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