嗜热烟曲霉SOD的性质表征及对毕赤酵母菌耐受性的影响  

Characterization of a Thermostable Superoxide Dismutase form Aspergillus fumigatus and Its Influence on Multi-stress Tolerance in Pichia pastoris

作　　者：崔保宇 卢海强[1] 王玉印 刘琼 田洪涛[1] 谷新晰[1] CUI Baoyu;LU Haiqiang;WANG Yuyin;LIU Qiong;TIAN Hongtao;GU Xinxi(College of Food Science and Technology,Hebei Agricultural University,Baoding 071001,Heibei;Beijing Shengtoda Biotechnology Co.,Beijing 100193)

机构地区：[1]河北农业大学食品科技学院,河北保定071001 [2]北京盛拓达生物技术有限公司,北京100193

出　　处：《中国食品学报》2025年第1期51-60,共10页Journal of Chinese Institute Of Food Science and Technology

基　　金：国家自然科学基金青年科学基金项目(31901631)。

摘　　要：基于毕赤酵母的密码子偏好性及高效表达规律,对嗜热烟曲霉来源的超氧化物歧化酶afSOD基因进行优化,使GC含量由59%降至41%,CAI值由0.69优化为0.96,优化前、后基因序列一致性为73.8%。经测定,重组菌株诱导72 h,酶活力可达213.78 U/mL。该重组酶AfSOD在pH 8.0和40℃条件下,具有最高催化活力,在中性环境和40~50℃稳定。金属离子Cu^(2+)对该酶活性的促进作用尤为显著,5 mmol/L的Cu^(2+)酶活提高80%,而Fe^(2+)则表现出明显的抑制效应。重组酶AfSOD对DPPH和羟自由基具有明显的清除作用,分别达到(97.65±1.46)%和(89.48±2.53)%。此外,重组酶AfSOD的高效表达可显著增强酵母菌株对温度、渗透压、盐、乙酸和乙醇的耐受性。本研究不仅为超氧化物歧化酶高效制备提供了一定的技术支撑,也为增强酵母菌生产耐受性提供理论参考。Superoxide dismutase is a cellular antioxidant with promising applications in food,cosmetic and pharmaceutical industries.In this study,based on the codon preference and efficient expression pattern of Pichia pastoris,the superoxide dismutase afSOD gene from Aspergillus fumigatus was optimized to reduce the GC content from 59%to 41%,and the CAI value was optimized from 0.69 to 0.96,and the gene sequence identity before and after the optimization was 73.8%.The recombinant strain induced an enzyme activity of up to 213.78 U/mL for 72 h.The recombinant enzyme AfSOD had the highest catalytic activity at pH 8.0 and 40℃,and was stable in a neutral environment and at 40-50℃.The promotion of the enzyme activity by the metal ion Cu^(2+)was particularly significant,with 5 mmol/L of Cu^(2+)increasing the enzyme activity by 80%,whereas Fe^(2+)showed a significant inhibitory effect.The recombinant enzyme AfSOD showed a significant scavenging effect on DPPH and hydroxyl radicals up to(97.65±1.46)%and(89.48±2.53)%.In addition,the efficient expression of recombinant enzyme AfSOD significantly enhanced the tolerance of yeast strains to temperature,osmotic pressure,salt,acetic acid and ethanol.This study not only provides some technical support for the efficient preparation of superoxide dismutase,but also provides theoretical support for the enhancement of yeast production tolerance.

关 键 词：超氧化物歧化酶 密码子优化 高效表达 酶学性质 耐受性 

分 类 号：TQ925[轻工技术与工程—发酵工程]