| 黄亚成,何斌,胡晓莎,苏龙兴,罗炳,刘林娅.刺梨果实转化酶提取方法的比较及酶学特性分析[J].中国南方果树,2024,53(4): |
| 刺梨果实转化酶提取方法的比较及酶学特性分析 |
| Different Extraction Methods and Enzymatic Characteristics of Invertase from the fruit of Rosa roxburghii |
| 投稿时间：2023-07-28 修订日期：2023-10-06 |
| DOI：10.13938/j.issn.1007-1431.20230345 |
| 中文关键词: 刺梨果实 转化酶 提取方法 酶学特性 |
| 英文关键词:Rosa roxburghii fruit invertase extraction methods enzymatic properties |
| 基金项目: |
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| 中文摘要: |
| 刺梨(Rosa roxburghii Tratt.)果实蔗糖转化酶通过调控蔗糖分解影响果实的生长发育和品质形成。基于不同提取方法能够影响植物蔗糖转化酶的提取效率和活性，本研究以‘贵农7号’刺梨为材料，比较了液氮研磨样品和提取液研磨样品下6种提取刺梨果实转化酶方法的效果，探究了可溶性转化酶的酶学特性。结果表明：液氮研磨法对刺梨果实转化酶活性影响较大，其中可溶性酸性转化酶（Soluable acid invertase，SAI）活性丧失严重，而细胞壁酸性转化酶(Cell wall-bound invertase，CWI)和中碱性转化酶(neutral or alkaline invertaes，N/A-INV)均无活性。采用提取液研磨样品，用Hepes-NaOH法提取刺梨果实转化酶的效果最好，SAI、CWI和N/A-INV的酶活性最高，分别为3.16、2.09和2.09μmol(glucose,fructose)/[g(FW)·h]；磷酸钠法次之。从比活力来看，Hepes-NaOH法提取的可溶性转化酶比活力亦最好。酶学特性分析表明SAI的最适温度为60℃、最适pH值为4.5-5.0、Km值为87.15mM；N/A-INV的最适温度为45-50℃、最适pH值为8.5、Km值为3.46mM。本研究结果为刺梨果实蔗糖转化酶的提取和蔗糖转化酶生物学功能的深入研究奠定了基础。 |
| 英文摘要: |
| Sucrose invertase is involved in the regulation of fruit development and formation of fruit quality via promote sucrose decomposition in Rosa roxburghii. Different extraction methods could affect the extraction efficiency and activity of plant sucrose invertase. In the study, ‘Guinong 5’ Rosa roxburghii was used as the material, six methods were investigated in extracting invertase from the fruit of Rosa roxburghii by two ways of grinding fruits: liquid nitrogen and extraction buffer, and the enzymatic characteristics of soluble invertase were explored. The result showed that the activities of invertases were seriously destroyed by grinding fruit in liquid nitrogen, especially those of cell wall-bound invertase (CWI) and neutral/alkaline invertase (N/A-INV) which were totally inactivated. After the fruits were ground in extraction buffers, the activity of crude soluble invertase extracted by Hepes-NaOH method revealed to be the best. The enzyme activities of SAI, CWI and N/A-INV were the highest, which were 3.16, 2.09 and 2.09μmol(glucose,fructose)/[g(FW)·h]. Phosphate-Na method is the second. With regard to specific enzymatic activity, the Hepes-NaOH method was the most appropriate.
SAI showed the highest enzymatic activity at pH 4.5-5.0 and 60℃, and had a Km of 87.15 mM for its substrate of sucrose. N/A-INV showed the highest enzymatic activity at pH 8.5 and 45-50℃, and had a Km of 3.46 mM for its substrate of sucrose. The results provide a foundation for the extraction of sucrose invertase and the further study of the biological function of sucrose invertase. |
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