The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, People's Republic of China.
College of Environmental Engineering, Nanjing Institute of Technology, Nanjing, 211167, People's Republic of China.
Appl Biochem Biotechnol. 2020 Nov;192(3):861-880. doi: 10.1007/s12010-020-03377-9. Epub 2020 Jul 1.
In this study, horseradish peroxidase C1A (HRP C1A) from Armoracia rusticana was expressed in Escherichia coli as an inclusion body. Subsequently, an active recombinant HRP C1A was obtained by refolding gradually using dilution-ultrafiltration. The recombinant HRP C1A was immobilized on agarose-chitosan hydrogel at 86.9 ± 2.5% of immobilization efficiency. After immobilization of the recombinant HRP C1A, the pH and temperature stability were improved and the reusability of the recombinant HPR C1A was achieved. The immobilized HRP C1A activity was retained above 80% after 6 cycles. The immobilized recombinant HRP C1A was used for the decolorization of four various dyes, including acid blue 129 (AB129), methyl blue (MB), methyl red (MR), and trypan blue (TB). The decolorization rates are all more than 70%, among which the decolorization effect of AB129 was the most significant (the decolorization rate was 76.3 ± 1.6%). Furthermore, a plausible decolorization pathway for AB129 was proposed based on the identified intermediates by ultraperformance liquid chromatography coupled with mass spectrometry (UPLC-MS). This is the first report of the putative mechanism on the decolorization of AB129 by HRP.
在这项研究中，辣根过氧化物酶 C1A（HRP C1A）来自芜菁被表达为大肠杆菌中的包涵体。随后，通过逐渐稀释-超滤复性获得了活性的重组 HRP C1A。将重组 HRP C1A 固定在琼脂糖-壳聚糖水凝胶上，固定效率为 86.9±2.5%。重组 HRP C1A 固定化后，提高了 pH 和温度稳定性，并实现了重组 HPR C1A 的可重复使用性。固定化的 HRP C1A 在 6 个循环后保留了超过 80%的活性。固定化的重组 HRP C1A 用于四种不同染料的脱色，包括酸性蓝 129（AB129）、甲基蓝（MB）、甲基红（MR）和锥虫蓝（TB）。脱色率均在 70%以上，其中 AB129 的脱色效果最为显著（脱色率为 76.3±1.6%）。此外，根据超高效液相色谱-质谱联用（UPLC-MS）鉴定的中间产物，提出了 AB129 可能的脱色途径。这是 HRP 对 AB129 脱色的作用机制的首次报道。
