Isolation, purification and characterization of a protease from the seeds of Artocarpus heterophyllus

IF 1.5 4区生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Indian journal of biochemistry & biophysics Pub Date : 2023-01-01 DOI:10.56042/ijbb.v60i9.4053

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Abstract

Proteases are being widely used in various industries like detergent, leather, food and pharmaceuticals.Protease was purified to homogeneity from the seeds of Artocarpus heterophyllus. The enzyme was found to be a tetramer having molecular mass of 74 kDa. Gelatin zymography showed a clear band of proteolysis. The enzyme isolated and purified was a serine protease, as indicated by its inhibition with PMSF. The enzyme was stable at broad pH and temperature ranges with pH and temperature optima at 8.5 and 50°C, respectively. The presence of some divalent ions enhanced the activity. With the addition of calcium, change in absorption and emission spectra was observed in spectrofluorometric analysis. The Km and Vmax for the enzyme was found to be 0.229 µM and 0.014 µM min-1, respectively, using BAPNA as a substrate. The enzyme consisted 4.44% alpha helix and 44.17% beta sheets when measured by CD spectra. Dynamic light scattering of the protease for particle size distribution revealed the mono-dispersity of the sample. Easy purification and paramount stability of protease makes it a good candidate for industrial and pharmaceutical applications.

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异叶树种子中一种蛋白酶的分离纯化及特性研究

蛋白酶被广泛应用于洗涤剂、皮革、食品和制药等各个行业。从异叶石竹种子中纯化出蛋白酶。该酶为四聚体，分子量为74 kDa。明胶酶谱图显示有清晰的蛋白水解带。分离纯化的酶为丝氨酸蛋白酶，对PMSF有抑制作用。该酶在较宽的pH和温度范围内稳定，pH和温度分别在8.5°C和50°C时最优。一些二价离子的存在增强了活性。在荧光光谱分析中，随着钙的加入，观察到吸收光谱和发射光谱的变化。以BAPNA为底物时，酶的Km和Vmax分别为0.229µM和0.014µM min-1。经CD谱测定，该酶由4.44%的α螺旋和44.17%的β螺旋组成。动态光散射的蛋白酶粒度分布揭示了样品的单分散性。蛋白酶的易纯化和极高的稳定性使其成为工业和制药应用的良好候选者。

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来源期刊

Indian journal of biochemistry & biophysics 生物-生化与分子生物学

CiteScore

2.90

自引率

50.00%

发文量

审稿时长

3 months

期刊介绍： Started in 1964, this journal publishes original research articles in the following areas: structure-function relationships of biomolecules; biomolecular recognition, protein-protein and protein-DNA interactions; gene-cloning, genetic engineering, genome analysis, gene targeting, gene expression, vectors, gene therapy; drug targeting, drug design; molecular basis of genetic diseases; conformational studies, computer simulation, novel DNA structures and their biological implications, protein folding; enzymes structure, catalytic mechanisms, regulation; membrane biochemistry, transport, ion channels, signal transduction, cell-cell communication, glycobiology; receptors, antigen-antibody binding, neurochemistry, ageing, apoptosis, cell cycle control; hormones, growth factors; oncogenes, host-virus interactions, viral assembly and structure; intermediary metabolism, molecular basis of disease processes, vitamins, coenzymes, carrier proteins, toxicology; plant and microbial biochemistry; surface forces, micelles and microemulsions, colloids, electrical phenomena, etc. in biological systems. Solicited peer reviewed articles on contemporary Themes and Methods in Biochemistry and Biophysics form an important feature of IJBB. Review articles on a current topic in the above fields are also considered. They must dwell more on research work done during the last couple of years in the field and authors should integrate their own work with that of others with acumen and authenticity, mere compilation of references by a third party is discouraged. While IJBB strongly promotes innovative novel research works for publication as full length papers, it also considers research data emanating from limited objectives, and extension of ongoing experimental works as ‘Notes’. IJBB follows “Double Blind Review process” where author names, affiliations and other correspondence details are removed to ensure fare evaluation. At the same time, reviewer names are not disclosed to authors.