蛋白质提取分离分析及米氏常数测定的实践及效果

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中图分类号 G642.1 文献标识码A 文章编号 1007-7731（2025）13-0118-04

DOI号10.16377/j.cnki.issn1007-7731.2025.13.030

Practice and effect of protein extraction， separation， analysis， and Michaelis constant determination

JIA Haoran HAN Peng ZHANG Yantao ZHANG Liquan （CollegeofLife Sciences，Inner Mongolia University，HohhotO1oO21， China）

AbstractProtein extraction，separation，analysis and determination of Michaelis constant are the comprehensive experimental coursecontents inthe teaching sylabusofbiochemistryexperiments.In thisexperimental practice， commercially available mung bean sprout stem segments were used as experimental materials.The total protein was extracted bygrinding method，andtheconcentrationand distributionof total protein were determinedby BCA method and SDS-PAGE electrophoresis.The Michaelis constant of acid phosphatase in total protein was determined using disodium p-nitrophenyl phosphate （PNPP-N ）as the substrate under acidic conditions[37 °C ，0.2 mol/L HAc-NaAc α bufr （pH=5.6）.The results showed that the total protein concentration of mung bean sprout stem segments was 4.374 mg/mL.Under acidic conditions，the Km of acid phosphatase in the total protein was 5.552 5 mmol/L，and the Vm was 0.0743mmol/（L·min）.This practical exercise significantly enhancedstudents’operational understandingof fundamentalbiochemicaltechniques including protein extraction/purification，quantitative analysis，and determination of Michaelis constant.Furthermore，it provided hands-on experience with esential methodologies such asprotein isolationand SDS-PAGE electrophoresis， while efectively fostering scientific curiosityand research enthusiasm among participants.

Keywords Biochemistry; extraction and determination of protein; Michaelis constant; mung bean sprouts

蛋白质是高校生物化学课程的核心教学内容之一。（剩余5522字）