甘露聚糖是自然界中半纤维素的第二大组分，广泛存在于饲料原料中，降低饲料营养物质的消化利用率。低温甘露聚糖酶在低温环境下（<30℃）高效水解甘露聚糖，可应用于低温环境和低温下的食品发酵和加工过程（如水产饲用酶制剂）。对低温甘露聚糖酶低温催化机理的研究可用于指导酶的分子改良，使不具低温活性的酶具有低温活性，扩展酶的应用范围。然而，国内外对低温甘露聚糖酶的报导极少，对其低温活性机理则尚未报导。云南高寒环境极可能孕育了新颖的低温微生物和低温甘露聚糖酶，具有重要的开发价值。本项目结合微生物的可培养法和直接克隆宏基因组中目的基因的m-TAIL-PCR方法，克隆云南高寒环境中的低温甘露聚糖酶，再通过点突变和肽段突变的方法揭示其低温活性机理。该项目对挖掘云南高寒环境中蕴藏的具有水产饲用潜力的新型低温甘露聚糖酶和揭示低温甘露聚糖酶的低温活性机理具有重要意义。

As the second most abundant hemicellulose in nature, mannans and heteromannans are widely distributed in animal feeds and reduce the digestion and absorption of nutrition. Effective breakdown of mannans and heteromannans at low temperatures (<30℃) mainly needs low-temperature-active endo-1,4- β -mannanases. The potential of low-temperature-active endo-1,4-β-mannanases include applications in low-temperature environments and biotechnological processes, such as aquaculture where low temperatures are required. Research on the mechanism of enzymatic adaptation to low temperatures is also important to improve endo-1,4-β-mannanases (gaining low-temperature activity) based on rational engineering. However, low-temperature-active endo-1,4-β-mannanases have rarely been reported, not to mention their catalytic mechanisms at low temperatures. Novel low-temperature-adapted microorganisms and low-temperature-active endo-1,4-β-mannanases, which have been forced to adapt to alpine environments in Yunnan province, are important to investigate. In this study, low-temperature-active endo-1,4-β-mannanases will be cloned from alpine environments in Yunnan province using both the classical culture-dependent and the culture-independent methods, called m-TAIL-PCR isolating functional genes directly from metagenome. Further amino acid residues and fragments mutageneses will be used to reveal the low-temperature-active mechanism of endo-1,4-β-mannanases. In a word, this study will contribute greatly to the study of low-temperature-active endo-1,4-β-mannanases in the following two aspects: screening novel low-temperature-active endo-1,4-β-mannanases with potential application in aquaculture from alpine environments in Yunnan province and revealing low-temperature-active mechanism of endo-1,4-β-mannanases.