关键词: 贻贝粘蛋白, 基因重组, 生物材料, 表征, 功效评价

Abstract:

In order to promote the application of recombinant mussel adhesive protein in the medical and cosmetics field， the recombinant mussel adhesive protein obtained from scale fermentation and purification of Escherichia coli was characterized and its efficacy was evaluated. Amino acid N-terminal sequencing， relative molecular weight analysis， amino acid composition analysis， protein purity analysis， endotoxin content， dihydroxyphenylalanine（DOPA） content of recombinant mussel adhesive protein were determined by the following methods： Edman degradation， matrix-assisted laser desorption ionization time-of-flight mass spectrometry （MALDI-TOF-MS）， phenyl-isothiocyanate （PITC）， nonreductive SDS-polyacrylamide gel electrophoresis （SDS-PAGE）， gel method， modified Arnow. The efficacy of recombinant mussel adhesive protein was evaluated by cell migration and repairing effect of zebrafish tail fin. Results showed that the obtained recombinant mussel adhesive protein was confirmed to be consistent with the theoretical primary structure， protein purity of more than 95%， endotoxin <10 EU·mg^-1， DOPA content above 5%. When the recombinant mussel adhesive protein concentration was 60 μg·mL^-1， the effect of promoting cell proliferation was the most obvious， and it had very significant activity （P<0.01）. The caudal fin area of zebrafish in sample group was significantly increased compared with model control group （P<0.001）. The results indicated that recombinant mussel adhesive protein can promote cell migration and repair healing and has the potential to be used as biomedical materials.

Key words: mussel adhesive protein, gene recombination, biological materials, representation, efficacy evaluation