利用重新利用的肌红蛋白开发维生素C生物传感器

Q2 Environmental Science
Sriyani Liyanage , Sriparna Ray
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引用次数: 0

摘要

铁基金属蛋白,肌红蛋白(Mb)不具有任何催化性质。它的基本功能是在脊椎动物体内储存氧气。通过修饰蛋白质活性位点附近的必需氨基酸,可以通过基因工程在Mb中产生过氧化物酶活性。这种“重新利用”的活性可以通过用非天然氨基酸(UAA)改变必需氨基酸来结合。此外,它易于表达和纯化,从而增加了其在催化和生物传感方面的应用。在这里,我们报道了用二羟基- l -苯丙氨酸(DOPA)对第29位亮氨酸残基进行遗传修饰,这是一种UAA,然后用甘氨酸取代H64。这种双突变使L29-DOPA/H64G突变体能够有效地催化硫代苯甲醚与H2O2的氧化,这是测试过氧化物酶活性的常用方法。事实上,在单氧化亚砜产物的同时,还产生了不同寻常的自由基反应产物——联苯、二苯硫醚等。这种独特的“重新利用”的部分可以用来设计生物传感器。这使得绿色化学第11条原则得以实现,该原则表达了实时监测不同物质的必要性。转基因和容易获得L29DOPA H64G Mb可以固定到电极基团,这可以很容易地检测electrocatalytic氧化的维生素C,因此,非盟/ L-Cys L29DOPAH64G突变Mb电极修改成抗坏血酸的生物传感器,它提供了一种方便的方式来监控有机食品含有维生素C L29DOPA / H64G Mb固定电极可以检测到低至5更易与维生素C / L,使用扫描率从50到200 mVs−1。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Utilizing repurposed myoglobin to develop biosensors of vitamin C
The iron-based metalloprotein, Myoglobin (Mb) does not possess any catalytic property. Its essential function is oxygen storage in vertebrates. By modifying the essential amino acids present near the active site of the protein, it is possible to generate peroxidase activity in Mb through genetic engineering. Such “repurposed” activity can be incorporated by altering the essential amino acid with an unnatural amino acid (UAA). In addition, it can be easily expressed and purified, thereby increasing its ease of application in catalysis and biosensing. Here, we report a genetic modification of the leucine residue at the 29th position with dihydroxy-L-phenylalanine (DOPA), which is an UAA, followed by the substitution of H64 by glycine. This double mutation enabled the L29-DOPA/H64G mutant to efficiently catalyze the oxidation of thioanisole with H2O2, a common method to test peroxidase activity. In fact, unusual products arising from radical mechanism – biphenyl, diphenyl sulfide, etc., were obtained along with mono-oxidized sulfoxide product. Such a uniquely “repurposed” moiety could be utilized to design biosensors. This enabled the achievement of the 11th principle of Green Chemistry, which expresses the need of monitoring different substances on a real-time basis. The genetically modified and easily procured L29DOPA H64G Mb could be immobilized onto a surface-modified electrode and this could easily detect the electrocatalytic oxidation of vitamin C. Thus, the Au/L-Cys/L29DOPAH64G mutant Mb electrode was modified into a biosensor for ascorbic acid, which provides a convenient way to monitor organic food items containing vitamin C. The L29DOPA/H64G Mb immobilized electrode could detect as low as 5 mmol/L of vitamin C, using scan rates from 50 to 200 mVs−1.
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来源期刊
Environmental Challenges
Environmental Challenges Environmental Science-Environmental Engineering
CiteScore
8.00
自引率
0.00%
发文量
249
审稿时长
8 weeks
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