Université de Toulouse, INSA, UPS, INP; LISBP, 31077 Toulouse, France.
J Ind Microbiol Biotechnol. 2012 Nov;39(11):1569-76. doi: 10.1007/s10295-012-1166-1. Epub 2012 Jul 10.
The secretome of Penicillium funiculosum contains two family GH7 enzymes, one of which (designated XynA) has been described as a xylanase. This is unusual because it is the only xylanase in family GH7, which is mainly composed of cellobiohydrolases and endoglucanases, and also because XynA is highly similar to the cellobiohydrolase I from Talaromyces emersonii and Trichoderma reesei (72 and 65 % identity, respectively). To probe this enigma, we investigated the biochemical properties of XynA, notably its activity on xylans and β-D-glucans. A highly pure sample of XynA was obtained and used to perform hydrolysis tests on polysaccharides. These revealed that XynA is 100-fold more active on β-1,4-glucan than on xylan. Likewise, XynA was active on both 4-nitrophenyl-β-D-lactopyranoside (pNP-β-D-Lac) and 4-nitrophenyl-β-D-cellobioside (pNP-cellobiose), which shows that XynA is principally an exo-acting type 1 cellobiohydrolase enzyme that displays 5.2-fold higher performance on pNP-cellobiose than on pNP-β-D-Lac. Finally, analyses performed using cellodextrins as substrate revealed that XynA mainly produced cellobiose (C2) from substrates containing three or more glucosyl subunits, and that C2 inhibits XynA at high concentrations (IC(50) (C2) = 17.7 μM). Overall, this study revealed that XynA displays typical cellobiohydrolase 1 activity and confirms that the description of this enzyme in public databases should be definitively amended. Moreover, the data provided here complete the information provided by a previous proteomics investigation and reveal that P. funiculosum secretes a complete set of cellulose-degrading enzymes.
木霉内切葡聚糖酶的分泌组包含两个 GH7 家族酶，其中一个（命名为 XynA）已被描述为木聚糖酶。这很不寻常，因为它是 GH7 家族中唯一的木聚糖酶，该家族主要由纤维二糖水解酶和内切葡聚糖酶组成，而且 XynA 与拟康氏木霉和里氏木霉的纤维二糖水解酶 I 非常相似（分别为 72%和 65%的同一性）。为了解开这个谜团，我们研究了 XynA 的生化特性，特别是它对木聚糖和β-D-葡聚糖的活性。我们获得了高度纯的 XynA 样品，并将其用于多糖水解试验。结果表明，XynA 对β-1,4-葡聚糖的活性比木聚糖高 100 倍。同样，XynA 对 4-硝基苯基-β-D-乳糖苷（pNP-β-D-Lac）和 4-硝基苯基-β-D-纤维二糖苷（pNP-纤维二糖）均具有活性，这表明 XynA 主要是一种外切作用的 1 型纤维二糖水解酶，对 pNP-纤维二糖的活性比 pNP-β-D-Lac 高 5.2 倍。最后，使用纤维二糖作为底物进行的分析表明，XynA 主要从含有三个或更多葡萄糖基单元的底物中产生纤维二糖（C2），并且 C2 在高浓度下抑制 XynA（IC50（C2）=17.7 μM）。总的来说，这项研究表明 XynA 表现出典型的纤维二糖水解酶 1 的活性，并证实了该酶在公共数据库中的描述应明确修正。此外，这里提供的数据补充了先前蛋白质组学研究提供的信息，并表明木霉分泌了一整套纤维素降解酶。
