首页|ATP含量与外源酶添加对罗非鱼肌原纤维蛋白磷酸化水平的影响

ATP含量与外源酶添加对罗非鱼肌原纤维蛋白磷酸化水平的影响

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蛋白质磷酸化修饰可影响肌肉品质，为探究外源酶添加对罗非鱼肌原纤维蛋白蛋白质磷酸化水平的影响，在向罗非鱼肌原纤维蛋白溶液中分别加入蛋白激酶A(protein kinase A，PKA)和碱性磷酸酶(alkaline phosphatase，AP)后进行体外孵育，通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和荧光染色测定不同时间段内的磷酸化水平变化。并通过将罗非鱼肌肉浸泡在不同浓度ATP溶液中测定肌原纤维蛋白的蛋白磷酸化水平以探究ATP对其的影响。结果显示，在0～72h，PKA组磷酸化水平均显著高于对照组和AP组，PKA组整体磷酸化水平从0h的0。35±0。01上升至12 h的0。37±0。01，而后下降至72 h的0。29±0。01，呈先上升后下降的趋势，其中，肌球蛋白重链磷酸化水平和肌动蛋白磷酸化水平分别从0h的0。73±0。01、0。86±0。01 下降至 72 h 的 0。58±0。02 和 0。68±0。01。当孵育时间为 0、4、24 和48h时，3组磷酸化水平均具有显著差异。在外源添加0。3 mol/L ATP后，结果显示肌原纤维蛋白整体磷酸化水平(0。46±0。00)显著高于对照组(0。42±0。01)。PKA可促进罗非鱼肌原纤维蛋白磷酸化修饰，AP则使其去磷酸化。研究表明，宰杀后罗非鱼肌肉中的ATP含量、PKA及AP活性水平是影响蛋白质磷酸化水平的关键因素。本研究可为探明罗非鱼品质变化机制与调控策略提供理论依据。
Effects of ATP content and exogenous enzyme addition on the phosphorylation level of myofibrillar protein in tilapia
Protein phosphorylation modification can affect muscle quality.In order to investigate the effect of exo-genous enzyme addition on the protein phosphorylation level of tilapia myofibrillar protein,tilapia myofibrillar protein solution was incubated in vitro by adding protein kinase A(PKA)and alkaline phosphatase(AP),respect-ively,and changes in phosphorylation level were determined by dodecyl sodium sulfate-polyacrylamide gel elec-trophoresis and fluorescence staining at different time intervals.In addition,the effect of adenosine triphosphate(ATP)on tilapia muscle was investigated by immersing the muscle in different concentrations of ATP solution to determine the level of myofibrillar fibrillar protein phosphorylation.The results showed that from 0 to 72 h,the phosphorylation level of PKA group was significantly higher than that of control group and AP group,and the overall phosphorylation level of PKA group increased from 0.35±0.01 at 0 h to 0.37±0.01 at 12 h,and then decreased to 0.29±0.01 at 72 h,with a general trend of increasing and then decreasing.In addition,the phos-phorylation level of myosin heavy chain and actin phosphorylation level decreased from 0.73±0.01 and 0.86±0.01 at 0 h to 0.58±0.02 and 0.68±0.01 at 72 h,respectively(P＜0.05).Phosphorylation levels were significantly differ-ent in all three groups when incubation time was 0,4,24 and 48 h.After addition of 0.3 mol/L exogenous ATP,the results showed that the overall phosphorylation level of myofibrillar proteins(0.46±0.001)was significantly higher than that of the control group(0.42±0.01).PKA promotes phosphorylation modification of myofibrillar proteins in tilapia,while alkaline phosphatase dephosphorylates them.It was shown that ATP content,PKA and AP activity levels in the muscle of slaughtered tilapia were key factors affecting the level of protein phosphorylation.This study may provide a theoretical basis for exploring the mechanism of quality change and regulatory strategies in tilapia.

tilapiamyofibrillar proteinprotein phosphorylationprotein kinase Aalkaline phosphataseATP

喻叶、魏涯、陈胜军、黄卉、岑剑伟、潘创、李春生、王迪、王悦齐、冯阳、赵永强

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中国水产科学研究院南海水产研究所,农业农村部水产品加工重点实验室,广东广州 510300

上海海洋大学食品学院,上海 201306

三亚热带水产研究院,海南省深远海渔业资源高效利用与加工重点实验室,海南三亚 572000

罗非鱼 肌原纤维蛋白 蛋白质磷酸化 蛋白激酶A 碱性磷酸酶 ATP

国家自然科学基金国家现代农业产业技术体系建设专项海南省自然科学基金海南省科技计划三亚崖州湾科技城联合项目广东省省级科技计划项目中国水产科学研究院中央级公益性科研院所基本科研业务费专项

32372257CARS-46323MS1222021CXLH00062023B02020100172023TD78

2024

10.11964/jfc.20240514515

水产学报
中国水产学会

水产学报

CSTPCD北大核心
影响因子：1.148
ISSN：1000-0615
年,卷(期)：2024.48(7)