肌动球蛋白磷酸化对其解离的影响

doi:10.7506/spkx1002-6630-201709004

食品科学 ›› 2017, Vol. 38 ›› Issue (9): 21-26.doi: 10.7506/spkx1002-6630-201709004

肌动球蛋白磷酸化对其解离的影响

高星，李欣，李铮，丁武，张德权

1.中国农业科学院农产品加工研究所，农业部农产品加工综合性重点实验室，北京 100193；2.西北农林科技大学食品科学与工程学院，陕西杨凌 712100

出版日期:2017-05-15 发布日期:2017-05-22

Effect of Phosphorylation on Actomyosin Dissociation

GAO Xing, LI Xin, LI Zheng, DING Wu, ZHANG Dequan

1. Key Laboratory of Agro-Products Processing, Ministry of Agriculture, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China; 2. College of Food Science and Engineering, Northwest A&F University, Yangling 712100, China

Online:2017-05-15 Published:2017-05-22

摘要/Abstract

摘要： 目的：研究改变肌动球蛋白粗提物磷酸化水平对其解离程度及ATPase活性的调控作用。方法：以肌动球蛋白粗提物为材料，添加蛋白激酶A（protein kinase A，PKA）和碱性磷酸酶（alkaline phosphatase，AP）改变蛋白的磷酸化水平（4 ℃孵育48 h），通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和荧光染色、蛋白质免疫印迹、ATPase活力测定试剂盒测定蛋白的磷酸化水平、游离肌动球蛋白含量和肌动球蛋白ATPase活力随孵育时间的变化。结果：PKA处理组蛋白样品中的肌钙蛋白T和肌球蛋白调节轻链的磷酸化水平显著高于对照组（P＜0.05），游离肌动蛋白含量显著高于对照组，且在0～2 h显著升高，2～48 h缓慢升高；肌动球蛋白ATPase活力显著低于对照组（P＜0.05），其活力随孵育时间延长显著升高（P＜0.05）；AP处理组的变化则与之相反。结论：肌钙蛋白T、肌球蛋白调节轻链的磷酸化降低了肌球蛋白与肌动蛋白之间的相互作用力（肌动球蛋白ATPase活力低），从而促进肌动球蛋白的解离。

关键词: 磷酸化, 肌球蛋白调节轻链, 肌钙蛋白T, 肌动球蛋白解离

Abstract: Objective: The objective of this study was to investigate the regulation of actomyosin dissociation and ATPase activity in crude actomyosin extract by changing the phosphorylation level of actomyosin. Methods: The crude actomyosin extract was incubated with protein kinase A and alkaline phosphatase at 4 ℃ for 48 h. Subsequently, the changes in protein phosphorylation, actomyosin dissociation and ATPase activity were measured by SDS-PAGE, Pro-Q Diamond phosphoprotein gel staining kit, Western blotting and ATPase activity assay kit. Results: The phosphorylation levels of troponin T (TnT) and myosin regulatory light chain (MRLC) were significantly higher in the protein kinase A group compared with the control group. The amount of liberated actin in the protein kinase A group was significantly higher than that in control group, and it increased significantly during the first two hours and increased slightly during the following 46 hours in both groups. In addition, the actomyosin ATPase activity decreased significantly in the protein kinase A group compared with the control group, and it increased significantly with incubation time in the two groups. However, the opposite changes were observed for the alkaline phosphatase group. Conclusion: The phosphorylation of TnT and MRLC reduces the binding force between myosin and actin and thereby promotes the dissociation of actomyosin.

Key words: phosphorylation, myosin regulatory light chain, troponin T, actomyosin dissociation

中图分类号:

TS201.2

高星，李欣，李铮，丁武，张德权. 肌动球蛋白磷酸化对其解离的影响[J]. 食品科学, 2017, 38(9): 21-26.

GAO Xing, LI Xin, LI Zheng, DING Wu, ZHANG Dequan. Effect of Phosphorylation on Actomyosin Dissociation[J]. FOOD SCIENCE, 2017, 38(9): 21-26.

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肌动球蛋白磷酸化对其解离的影响

Effect of Phosphorylation on Actomyosin Dissociation

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