Abstract：
Mussel foot proteins (Mfps) possess unique binding properties to various surfaces due to the presence of L-3,4-dihydroxyphenylalanine (DOPA). Mytilus edulis foot protein-3 (Mefp-3) is one of several proteins in the byssal adhesive plaque. Its localization at the plaque-substrate interface approved that Mefp-3 plays a key role in adhesion. Therefore, the protein is suitable for the development of innovative bio-based binders. However, recombinant Mfp-3s are mainly purified from inclusion bodies under denaturing conditions. Here, we describe a robust and reproducible protocol for obtaining soluble and tag-free Mefp-3 using the SUMO-fusion technology. Additionally, a microbial tyrosinase from Verrucomicrobium spinosum was used for the in vitro hydroxylation of peptide-bound tyrosines in Mefp-3 for the first time. The highly hydroxylated Mefp-3, confirmed by MALDI-TOF-MS, exhibited excellent adhesive properties comparable to a commercial glue. These results demonstrate a concerted and simplified high yield production process for recombinant soluble and tag-free Mfp3-based proteins with on demand DOPA modification.
摘要：
由于存在L-3,4-二羟基苯丙氨酸(DOPA),贻贝足蛋白(Mfps)对各种表面具有独特的结合性质。Mytilusedulis足蛋白3（Mefp-3）是底骨粘附斑块中的几种蛋白质之一。它在斑块-基质界面的定位证明Mefp-3在粘附中起关键作用。因此,该蛋白质适合开发创新的生物基粘合剂。然而,重组Mfp-3s主要在变性条件下从包涵体中纯化。这里,我们描述了使用SUMO融合技术获得可溶性和无标签的Mefp-3的稳健且可重复的方案.此外,来自马尾草的微生物酪氨酸酶首次用于Mefp-3中与肽结合的酪氨酸的体外羟基化。通过MALDI-TOF-MS确认的高度羟基化的Mefp-3表现出与商业胶相当的优异的粘合性能。这些结果证明了具有按需DOPA修饰的重组可溶性和无标签的基于Mfp3的蛋白质的一致和简化的高产率生产过程。
