Single-Step Purification of Catalase Enzyme From Human Blood Erythrocytes Using Affinity Chromatography Technique.

IF 2.6 3区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY BioMed Research International Pub Date : 2024-07-09 eCollection Date: 2024-01-01 DOI:10.1155/2024/2222098
Kübra Çıkrıkcı, Nahit Gencer
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Abstract

In this study, we aimed to isolate and purify catalase from human blood erythrocytes by using a newly synthesized affinity gel. The synthesized ω-amino hexyl agarose-1,2,3-triazole-5-carboxylic acid affinity gel was analyzed by FT-IR. Then, different buffer, pH, and ionic strength parameters were optimized to determine the equilibration, washing, and elution buffer conditions. The catalase was purified from human blood erythrocytes with a specific activity of 45.58 EU/mg, purification fold of 529.50, and a yield of 0.416% using the synthesized new affinity gel. The purity and molecular weight of the enzyme were analyzed by SDS-PAGE, and a single band at 60 kDa was observed for catalase. The optimum reaction temperature of the catalase was found to be 30°C, while the thermal stability temperature was 60°C. The Km and Vmax of the enzyme for hydrogen peroxide were calculated at 0.125 mM and 2500 U mL-1, respectively.

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利用亲和层析技术从人血红细胞中单步纯化过氧化氢酶
本研究旨在利用一种新合成的亲和凝胶从人血红细胞中分离纯化过氧化氢酶。通过傅立叶变换红外光谱分析了合成的ω-氨基己基琼脂糖-1,2,3-三唑-5-羧酸亲和凝胶。然后,对不同的缓冲液、pH 值和离子强度参数进行了优化,以确定平衡、洗涤和洗脱缓冲液的条件。利用合成的新型亲和凝胶从人血红细胞中纯化出过氧化氢酶,其比活度为 45.58 EU/mg,纯化倍数为 529.50,产率为 0.416%。用 SDS-PAGE 分析了酶的纯度和分子量,发现过氧化氢酶有一条 60 kDa 的条带。过氧化氢酶的最佳反应温度为 30°C,热稳定性温度为 60°C。计算得出该酶对过氧化氢的 Km 和 Vmax 分别为 0.125 mM 和 2500 U mL-1。
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来源期刊
BioMed Research International
BioMed Research International BIOTECHNOLOGY & APPLIED MICROBIOLOGY-MEDICINE, RESEARCH & EXPERIMENTAL
CiteScore
6.70
自引率
0.00%
发文量
1942
审稿时长
19 weeks
期刊介绍: BioMed Research International is a peer-reviewed, Open Access journal that publishes original research articles, review articles, and clinical studies covering a wide range of subjects in life sciences and medicine. The journal is divided into 55 subject areas.
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