重组贻贝粘蛋白Mfp-3P的制备及促进伤口愈合的作用

Preparation of recombinant mussel mucin Mfp-3P and its promotion of wound healing

摘要:

为了研究重组贻贝粘蛋白的制备方法及其在伤口愈合中的作用，本研究将贻贝的Mfp-3和preCol-P肽部分片段进行融合，以毕赤酵母为宿主微生物进行表达，经过发酵和纯化，获得了纯的重组贻贝粘蛋白产物，并在细胞水平检测其对小鼠成纤维细胞L929增殖和迁移的影响，以及在动物水平检测其在大鼠全层皮肤切除后促进伤口愈合的能力。结果表明，制备获得的重组贻贝粘蛋白纯度为96.49%，蛋白含量为90.28%，3,4-二羟基苯丙氨酸(3,4-dihydroxyphenylalanine, DOPA)含量为0.73 wt%，内毒素含量<0.5 EU/mg，具有显著地促进小鼠成纤维细胞迁移、增殖及促大鼠皮肤创面伤口愈合的作用。本研究结果表明，采用融合表达的思路能够实现贻贝Mfp-3在毕赤酵母表达体系的高效表达，其产物易于纯化，获取的纯品质量稳定，具有促进伤口愈合的作用，具备作为医用生物材料的潜质。

Abstract:

To investigate the role of recombinant mussel mucin in wound healing, we aimed to prepare this mucin using Pichia pastoris as the host microbe. Our method involved constructing a genetically engineered strain of P. pastoris that expressed a fusion protein consisting of Mfp-3 and preCol-P peptide segments of mussel. After fermentation and purification, we obtained a pure recombinant mussel mucin product. We then conducted experiments to evaluate its effect at both the cellular and animal levels. At the cellular level, we examined its impact on the proliferation and migration of mouse fibroblast L929. At the animal level, we assessed its ability to promote wound healing after full-layer skin resection in rats. Our results showed that the recombinant mussel mucin protein has a content of 90.28% and a purity of 96.49%. The content of 3,4-dihydroxyphenylalanine (DOPA) was 0.73 wt%, and the endotoxin content was less than 0.5 EU/mg. Importantly, the recombinant mussel mucin protein significantly promoted both the migration and proliferation of mouse fibroblast, as well as the wound healing in rat skin. In conclusion, our findings demonstrate that recombinant mussel mucin has the potential to promote wound healing and can be considered a promising medical biomaterial.