蛋白质磷酸化与泛素化互作调控肌间线蛋白降解的分子机理

嫩度是最重要的肉品品质，肌原纤维蛋白降解是宰后肌肉嫩化的本质，肌间线蛋白是维持肌纤维结构完整的关键肌原纤维蛋白，影响肉品嫩化进程。申请人研究发现，蛋白质磷酸化和泛素化修饰影响肌间线蛋白降解，但具体的影响机理尚不清楚。本项目以嫩度差异显著的肉样为试验材料，采用免疫印迹、Phos-tag SDS-PAGE电泳等技术，研究蛋白质磷酸化和泛素化与肌间线蛋白降解的表征关系；建立体外肌肉模型，通过外源调控磷酸化水平，研究蛋白质磷酸化对肌间线蛋白泛素化水平、位点、降解程度和降解产物的影响；通过外源调控泛素化水平，研究蛋白质泛素化对肌间线蛋白磷酸化水平、位点、降解程度和降解产物的影响；采用生物信息学、蛋白质结构模拟等技术，研究蛋白质磷酸化和泛素化互作对肌间线蛋白结构的影响，揭示蛋白质磷酸化和泛素化互作调控肌间线蛋白降解的分子机理，为完善和发展肉品嫩化理论、研发肉品精准嫩化技术提供依据。

Tenderness is the most important meat quality attribute, myofibril protein degradation is the essence of meat tenderization postmortem. Desmin is the key myofibril protein which keeps the integrity of muscle fiber and influences meat tenderness. The applicant found that protein phosphorylation and ubiquitination regulate desmin degradation, but the detailed mechanism is unknown. The objective of this study is to investigate the molecular mechanism of protein phosphorylation and ubiquitination cross talk regulating desmin degradation, in order to provide evidence for improvement of meat tenderization theory and development of precise meat tenderization technology. Meat sample with different tenderness will be obtained to study the correlation between protein phosphorylation, ubiquitination and desmin degradation using the technologies of immunoblot, Phos-tag SDS-PAGE, etc.; the effects of protein phosphorylation on desmin ubiquitination level and sites, degradation level and degradation products will be studied using in vitro muscle system through regulating phosphorylation level; the effects of protein ubiquitination on desmin phosphorylation level and sites, degradation level and degradation products will be studied using in vitro muscle system through regulating ubiquitination level; and the effects of protein phosphorylation and ubiquitination cross talk on desmin structure will be studied using bioinformatics and protein structure simulation program.