Abstract The formation of underwater adhesion is a complicated physiological process and many different types of enzymes are found to be essential apart from structural proteins. Previous studies have shown that various tyrosinases were present in marine adhesives, but little information is available about the over-expression and enzymatic characterization of these enzymes. Specifically, this study first identified four significantly up-regulated tyrosinases in the pedal disc of Haliplanella luciae by means of multi-omics technology, and made preliminary bioinformatics predictions. Sequence alignment showed that the Tyr1_Hl contained six conserved His residues that bind to copper ions, of which a tyrosinase with diphenolase activity named as Tyr1_HlΔ, was expressed in Escherichia coli BL21 (DE3) cells and purified by affinity chromatography. Enzymatic characterization showed that the activity of Tyr1_HlΔ was Cu2+ dependent and maximum catalytic activities were in 20 mM Tris–HCl (pH 8.0) at 37 °C. In summary, we identified novel tyrosinases in the pedal disks of sea anemone for the first time and the Tyr1_HlΔ was successfully recombinant expressed. Our study will provide basis for future exploration of bio-adhesion mechanism and design of bio-adhesives derived from sea anemones.

中文翻译：

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海葵足盘酪氨酸酶的发现与表征

摘要 水下粘附的形成是一个复杂的生理过程，除结构蛋白外，还发现许多不同类型的酶是必不可少的。以前的研究表明，海洋粘合剂中存在各种酪氨酸酶，但关于这些酶的过度表达和酶学表征的信息很少。具体而言，本研究首先利用多组学技术，在Haliplanella luciae足盘中鉴定出四种显着上调的酪氨酸酶，并进行了初步的生物信息学预测。序列比对表明，Tyr1_Hl 含有 6 个与铜离子结合的保守 His 残基，其中具有双酚酶活性的酪氨酸酶 Tyr1_HlΔ 在大肠杆菌BL21（DE3）细胞中表达，并通过亲和层析纯化。酶学表征表明，Tyr1_HlΔ 的活性依赖于 Cu2+，最大催化活性在 37°C 下的 20 mM Tris-HCl (pH 8.0) 中。总之，我们首次在海葵足盘中发现了新型酪氨酸酶，并成功重组表达了Tyr1_HlΔ。我们的研究将为今后探索海葵生物黏附机制和设计生物黏附剂提供依据。