Production, purification, and characterization of metalloprotease from Candida kefyr 41 PSB

详细信息    查看全文

• 作者：Sevgi Yavuz^a ; Samet Kocabay^b ; Serap Ç ; etinkaya^c ; Birnur Akkaya^c ; ^{bakkaya@cumhuriyet.edu.tr} ; ^{pergamonchem@gmail.com} ; Recep Akkaya^d ; Ali Fazil Yenidunya^c ; Mustafa Zahir Bakıcı^e
• 关键词：BSA ; bovine serum albumin ; CMC ; critical micelle concentration ; EDTA ; ethylenediamine tetraacetic acid ; DMSO ; dimethyl sulfoxide ; DTT ; dithiothreitol ; ICP/MS ; inductively coupled plasma mass spectrometry ; MALDI-TOF MS ; matrix-assisted laser desorption/ionization time-of-flight mass spectrometry ; MRS ; man ; rogosa and sharp medium ; SD ; standard deviation ; SDS-PAGE ; sodium dodesyl sulphat-polyacrylamide gel electrophoresis ; PMSF ; phenylmethylsulfonyl fluoride ; TCA ; trichloroacetic acid
• 刊名：International Journal of Biological Macromolecules
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：94
• 期：part_PA
• 页码：106-113
• 全文大小：1172 K
• 卷排序：94

文摘

A thermostable metalloprotease, produced from an environmental strain of Candida kefyr 41 PSB, was purified 16 fold with a 60% yield by cold ethanol precipitation and affinity chromatography (bentonite-acrylamide-cysteine microcomposite). The purified enzyme appeared as a single protein band at 43 kDa. Its optimum pH and temperature points were found to be 7.0 and 105 °C, respectively. Km and Vmax values of the enzyme were determined to be 3.5 mg/mL and 4.4 μmol mL−1 min−1, 1.65 mg/mL and 6.1 μmol mL−1 min−1, using casein and gelatine as the substrates, respectively. The activity was inhibited by using ethylenediamine tetraacetic acid (EDTA), indicating that the enzyme was a metalloprotease. Stability of the enzyme was investigated by using thermodynamic and kinetic parameters. The thermal inactivation profile of the enzyme conformed to the first order kinetics. The half life of the enzyme at 95, 105, 115, 125 and 135 °C was 1310, 610, 220, 150, and 86 min, respectively.