调控配位不饱和位点增强锆基金属有机框架纳米酶的蛋白水解酶活性及应用

Nanozymatic activity of Zr-based MOFs:Tuning the coordinatively unsaturated metal sites for enhancing hydrolytic activity toward peptide bonds

目的:开发高效稳定的蛋白水解酶纳米酶,研究配位不饱和位点与肽键水解活性的关系。方法:制备3种不同配位(12/6/4配位)不饱和位点的锆基金属有机框架纳米酶(Zr-MOFs纳米酶)调控蛋白水解酶活性。以双甘肽(Gly-Gly)水解率为指标,评估3种Zr-MOFs纳米酶的蛋白水解酶活性。随后,使用性能最优的蛋白水解酶纳米酶水解大豆蛋白、鱼糜蛋白和酪蛋白,利用SDS-聚丙烯酰胺凝胶电泳分离水解产物,通过考马斯亮蓝对分离的条带染色,考察人工蛋白酶对3种蛋白的水解效率。结果:在3种不同配位不饱和位点Zr-MOFs纳米酶中,6-配位Zr-MOF纳米酶的蛋白水解酶活性最高,水解率为52%。与非催化水解相比,水解反应速率提高了2.63×10^(3)倍。蛋白水解酶纳米酶可水解食品工业中3种常见的蛋白质,对鱼糜蛋白的水解效率最高。结论:通过调控Zr-MOFs纳米酶的配位不饱和位点可以增强蛋白水解酶活性。

Objective:To develop efficient and stable artificial proteases and investigate the relationship between coordinatively unsaturated metal sites and hydrolysis activity of peptide bond.Methods:Three zirconium-based metal-organic frameworks(Zr-MOFs)with different coordinatively unsaturated sites(12-connected Zr-MOF,6-connected Zr-MOF,and 4-connected Zr-MOF)were prepared to modulate protein hydrolase activity.The protein hydrolase activity of three Zr-MOFs was evaluated by the hydrolysis rate of bis-glycopeptide(Gly-Gly)as an indicator.Subsequently,soy protein,surimi protein and casein were hydrolyzed using the best performing protein hydrolase nanoenzymes,and the hydrolysis products were separated using SDS-polyacrylamide gel electrophoresis,and the hydrolysis efficiency of the three proteins by artificial proteases was investigated by staining the separated bands with Komas Brilliant Blue.Results:Among the three Zr-MOFs proteases,the 6-connected Zr-MOF showed the highest hydrolysis(52%)efficiency toward the Gly-Gly.The hydrolysis reaction rate was increased 2.63×10^(3) times compared the uncatalyzed hydrolysis.In addition,the 6-connected Zr-MOF could respectively hydrolyze three commonly proteins in the food industry and had the highest catalytic effect on surimi protein.Conclusion:These results revealed that the ability of Zr-MOF to hydrolyze peptide bonds could be enhanced by tuning the coordinatively unsaturated sites.The study provided a new idea for the application of artificial proteases in food industry.