Grenier J, Potvin C, Trudel J, Asselin A
Département de Phytologie, Faculté des sciences de l'agriculture et de l'alimentation, Université Laval, Québec, Canada.
Plant J. 1999 Aug;19(4):473-80. doi: 10.1046/j.1365-313x.1999.00551.x.
Thaumatin and 12 purified thaumatin-like (TL) proteins were surveyed for their capacity to hydrolyse beta-1,3-glucans by using an in-gel glucanase assay. Six TL proteins identified by N-terminal amino acid microsequencing were found to be active on carboxymethyl(CM)-pachyman: a barley leaf stress-related permatin, two tomato fruit osmotins, a cherry fruit and two tobacco stigma proteins. TL enzymes ranged in specific activity from 0.07 to 89 nkat mg-1 with CM-pachyman as substrate. Hydrolytic activities were not restricted to TL proteins strongly binding to water-insoluble beta-1,3-glucans since the two osmotins were active without tight binding to pachyman. Some TL proteins hydrolysed crude fungal walls and one barley TL enzyme even lysed fungal spores. No activity was observed on laminarin in the in-gel hydrolase assay. Thin-layer chromatography revealed that the six enzymes acted as endo-beta-1, 3-glucanases leading to the formation of various oligoglucosides. Thus far, the TL enzymes (EC 3.2.1.x) appeared different from the well-known beta-1,3-glucanases (EC 3.2.1.39). No activity was found with thaumatin, zeamatin, tobacco leaf PR-R protein and four stress-related TL proteins from barley and pea. This is the first demonstration that diverse TL proteins are enzymatically active. The functions of some TL proteins must be reassessed because they display endo-beta-1,3-glucanase activity on polymeric beta-1, 3-glucans.
通过凝胶内葡聚糖酶测定法，对奇异果甜蛋白和12种纯化的类奇异果甜蛋白（TL）进行了水解β-1,3-葡聚糖能力的检测。通过N端氨基酸微量测序鉴定的6种TL蛋白在羧甲基（CM）-茯苓聚糖上具有活性：一种大麦叶片胁迫相关的渗透素、两种番茄果实渗透素、一种樱桃果实和两种烟草柱头蛋白。以CM-茯苓聚糖为底物时，TL酶的比活性范围为0.07至89 nkat mg-1。水解活性并不局限于与水不溶性β-1,3-葡聚糖强烈结合的TL蛋白，因为这两种渗透素在未紧密结合茯苓聚糖的情况下也具有活性。一些TL蛋白能水解粗真菌细胞壁，一种大麦TL酶甚至能裂解真菌孢子。在凝胶内水解酶测定中未观察到对海带多糖的活性。薄层色谱显示，这六种酶作为内切β-1,3-葡聚糖酶，导致形成各种低聚糖苷。到目前为止，TL酶（EC 3.2.1.x）似乎与著名的β-1,3-葡聚糖酶（EC 3.2.1.39）不同。在奇异果甜蛋白、玉米甜蛋白、烟草叶片PR-R蛋白以及来自大麦和豌豆的四种胁迫相关TL蛋白中未发现活性。这是首次证明多种TL蛋白具有酶活性。一些TL蛋白的功能必须重新评估，因为它们对聚合β-1,3-葡聚糖显示出内切β-1,3-葡聚糖酶活性。
