耐低温纤维素降解真菌的筛选、鉴定及酶活性研究

Screening and Identification of Cold-adapted Fungi with Cellulose Degradation and Its Enzymatic Properties

  • 摘要:
    目的 为筛选低温下高效降解纤维素的菌种资源,促进秸秆在低温条件的快速腐解。
    方法 本研究利用低温稀释平板涂布法分离、刚果红定性、内切β-葡聚糖酶活力定量等分析方法进行低温纤维素降解真菌的筛选,依据菌株形态特征及rDNA-ITS序列分析鉴定菌种,探究其在不同温度下的产酶能力,并对粗酶提取物进行酶学性质的初步研究。
    结果 ① 筛选到一株低温下高效产纤维素酶的真菌ZY-50,鉴定为球芽异普可尼亚菌(Metapochonia-bulbillosa),目前尚无该菌产纤维素酶的报道;② 球芽异普可尼亚菌ZY-50可在低温条件下生长并具有较强产纤维素酶能力,其内切β-葡聚糖酶活力在28℃、16℃、10℃温度下分别为76.03 U mL−1、128.34 U mL−1和115.67 U mL−1,该菌在16℃下产外切β-葡聚糖酶和β-葡萄糖苷酶的能力较稳定,最高分别为2.31 U mL−1、2.43 U mL−1,16℃下滤纸酶活力最高为7.89 U mL−1;③粗酶提取物的内切β-葡聚糖酶最适作用温度为60℃,在10 ~ 40℃温度下,在1.5 h内相对酶活为90%,最适pH为5.0,10 mmol L−1 Co2 + 对其活性有显著的促进作用。
    结论 球芽异普可尼亚菌ZY-50可在低温条件下生长且产纤维素酶活力较强,在促进秸秆低温腐熟方面具有较好的的应用潜力。

     

    Abstract:
    Objective The aim was to screen strain resources for efficient cellulose degradation and to promote rapid decomposition of straw at low temperature.
    Method Low-temperature cellulose degrading fungi were screened by using low-temperature dilution plate coating method, Congo red characterization and quantification of endo-β-glucanase activities. The strains were identified based on morphological characteristics and rDNA-ITS sequence analysis, their enzyme production capacities at different temperatures were investigated, and preliminary studies on the enzymatic properties of crude enzyme extracts were conducted.
    Result ① A cold-adapted fungus, namely ZY-50, was obtained and identified as Metapochonia-bulbillosa. There are no reports of cellulase production by this fungus. ② Metapochonia-bulbillosa ZY-50 could grow at low temperatures and has strong cellulase production ability. Its Endo β-glucanase activities were 76.03 U mL−1, 128.34 U mL−1, and 115.67 U mL−1 at 28℃, 16℃ and 10℃ respectively, and the abilities of the fungus to produce Exo β-glucanase and β-Glucosidase were more stable at 16℃. The highest activities of them were 2.31 U mL−1 and 2.43 U mL−1. The synergistic effect of the three cellulases was stronger at 16℃, with the highest filter paper activity of 7.89 U mL−1. ③ The endo-β-glucanase of the crude enzyme extract had an optimum action temperature of 60℃. The relative enzyme activity was 90% within 1.5 h at temperatures between 10 and 40℃. The optimum pH was 5.0, and its activity was promoted by 10 mmol L−1 Co2 + .
    Conclusion Metapochonia-bulbillosa ZY-50 can grow at low temperature and has a strong cellulase production ability, which has a high potential to promote straw decomposition at low temperature.

     

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