Department of Biochemistry, Jawaharlal Nehru Medical College, A.M.U., Aligarh, India.
Department of Biochemistry, Jawaharlal Nehru Medical College, A.M.U., Aligarh, India.
Int J Biol Macromol. 2019 Apr 1;126:1201-1212. doi: 10.1016/j.ijbiomac.2019.01.018. Epub 2019 Jan 6.
Methylglyoxal (MGO) is a highly reactive α-oxoaldehyde. It reacts with basic amino acids of the proteins to form advanced glycation end products (AGEs). Fibrinogen is a soluble multi-domain glycoprotein whose major function is to form fibrin clots that prevent blood loss upon vascular injury. In the present study, fibrinogen was incubated with varying concentration of MGO for 7 days followed by its biochemical and biophysical analysis. Glycated plasma fibrinogen (MGO-fibrinogen); exhibited hyperchromicity, a drop in tryptophan and intrinsic fluorescence, augmented AGE-specific fluorescence and melting temperature. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) results showed decrease in mobility of MGO-fibrinogen. Structural perturbations in secondary and tertiary structure were identified by fourier transform-infrared spectroscopy (FT-IR), followed by far and near-UV circular dichroism (CD). Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF-TOF) mass spectrometry studies suggested increase in molecular mass of MGO-fibrinogen. Amyloid like aggregates were confirmed by Thioflavin T (ThT), Congo red assay (CR), scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The ketoamine moieties, carbonyl content (CO), hydroxymethylfurfural (HMF), superoxide and hydroxyl content were markedly elevated, whereas, total antioxidant capacity (TAC) and free thiol content decreased in MGO-fibrinogen as compared to the native protein. These investigations confirmed the structural and functional alterations in MGO-fibrinogen which leads to different physiological conditions like diabetes mellitus, cardiovascular disease etc.
甲基乙二醛（MGO）是一种高度反应性的α-氧醛。它与蛋白质的碱性氨基酸反应，形成晚期糖基化终产物（AGEs）。纤维蛋白原是一种可溶性的多结构域糖蛋白，其主要功能是形成纤维蛋白凝块，防止血管损伤时出血。在本研究中，纤维蛋白原与不同浓度的 MGO 孵育 7 天，然后进行生化和生物物理分析。糖基化血浆纤维蛋白原（MGO-纤维蛋白原）表现出增色、色氨酸和固有荧光下降、AGE 特异性荧光增强和熔点升高。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）结果表明 MGO-纤维蛋白原的迁移率降低。傅里叶变换红外光谱（FT-IR）分析表明二级和三级结构发生了结构扰动，随后进行了远和近紫外圆二色性（CD）分析。基质辅助激光解吸电离飞行时间（MALDI-TOF-TOF）质谱研究表明 MGO-纤维蛋白原的分子量增加。通过噻唑蓝（ThT）、刚果红测定（CR）、扫描电子显微镜（SEM）和透射电子显微镜（TEM）证实了类淀粉样聚集物的形成。酮胺部分、羰基含量（CO）、羟甲基糠醛（HMF）、超氧阴离子和羟基含量明显升高，而 MGO-纤维蛋白原的总抗氧化能力（TAC）和游离巯基含量则低于天然蛋白。这些研究证实了 MGO-纤维蛋白原的结构和功能改变，导致不同的生理状况，如糖尿病、心血管疾病等。
