Plant Physiology Lab, Taiwan Forestry Research Institute, 53 Nan-Hai Road, Taipei, Taiwan, ROC.
Plant Physiol. 1989 Dec;91(4):1445-53. doi: 10.1104/pp.91.4.1445.
Pectinmethylesterase from the pericarp of jelly fig (Ficus awkeotsang) achenes was extracted and purified to a specific activity of 289 micromole proton produced per minute per milligram protein. Pectinmethylesterase, a major protein with high specific activity in the crude extract, was monomeric with a molecular weight of 38,000. The enzyme preparation was stable in distilled water at 4 degrees C for at least 6 months, and at 60 degrees C for at least 10 minutes. This enzyme functioned optimally at pH 6.5 to 7.5 when the assay mixture contained no NaCl or at low NaCl concentration. The pH optimum shifted to lower pH as the NaCl concentration was increased. The K(m) value for pectin was 0.75 milligram per milliliter pectin, corresponding to a V(max) value of 310 micromoles per minute per milligram protein. Inhibition studies with antibodies indicated that jelly fig achene pectinmethylesterase and the two other pectinmethylesterases from orange and tomato were similar in their active site conformation; however, the surface determinants may be very different because no precipitation between anti-jelly fig pectinmethylesterase immune serum and the pectin methylesterase from orange and tomato could be observed in the double immunodiffusion analysis. Specific antisera raised against jelly fig achene pectinmethylesterase in a Western blot experiment also showed low similarity between jelly fig pectinmethylesterase with that from orange and tomato. This observation was also supported by the very low isoelectric point (pH 3.5) of jelly fig pectinmethylesterase, compared with high isoelectric points reported for most of the pectinmethylesterases. Amino acid composition and N-terminal sequence have been obtained. High homology of the N-terminal amino acid residues between jelly fig and tomato pectinmethylesterase (O Markovic, H Jornvall [1986] Eur J Biochem 158: 455-462) was observed. Pectinmethylesterase activity causes the release of protons from the deesterification of pectin such that a low pH environment is created, and this may be related to the cell growth. Pectinmethylesterase is not needed for jelly fig seed germination, however the gel formed from pectin and pectinmethylesterase may insure a water source for the germinating jelly fig seeds.
从木奶果（Ficus awkeotsang）种皮中提取和纯化果胶甲酯酶，使其比活达到每毫克蛋白每分钟产生 289 微摩尔质子。果胶甲酯酶是粗提物中具有高比活的主要蛋白质，分子量为 38000，为单体。酶制剂在 4°C 的蒸馏水中至少稳定 6 个月，在 60°C 下至少 10 分钟。该酶在不含 NaCl 或低浓度 NaCl 的测定混合物中，在 pH6.5 到 7.5 时最佳发挥作用。当 NaCl 浓度增加时，pH 最适值向更低 pH 移动。果胶的 K(m)值为 0.75 毫克/毫升果胶，对应的 V(max)值为 310 微摩尔/分钟/毫克蛋白。用抗体进行的抑制研究表明，木奶果种皮果胶甲酯酶与来自橙子和番茄的另外两种果胶甲酯酶在活性部位构象上相似；然而，表面决定因素可能非常不同，因为在双免疫扩散分析中，没有观察到抗木奶果果胶甲酯酶免疫血清与来自橙子和番茄的果胶甲酯酶之间发生沉淀。在 Western blot 实验中，针对木奶果种皮果胶甲酯酶产生的特异性抗血清也表明，木奶果果胶甲酯酶与来自橙子和番茄的果胶甲酯酶之间的相似性很低。这种观察结果也得到了木奶果果胶甲酯酶非常低的等电点（pH3.5）的支持，而大多数果胶甲酯酶的等电点报告值较高。已经获得了氨基酸组成和 N 端序列。木奶果和番茄果胶甲酯酶（O Markovic，H Jornvall [1986] Eur J Biochem 158：455-462）的 N 端氨基酸残基具有高度同源性。果胶甲酯酶活性导致果胶脱酯化释放质子，从而形成低 pH 环境，这可能与细胞生长有关。果胶甲酯酶不是木奶果种子萌发所必需的，然而果胶和果胶甲酯酶形成的凝胶可能为萌发的木奶果种子提供水分来源。
