摘要
裂解性多糖单加氧酶(LPMOs)辅助活性9家族蛋白(AA9)可有效促进纤维素酶降解纤维素。为研究N-糖基化对来源于黑曲霉的AA9蛋白AnLPMO15g与纤维素酶协同性的影响,采用定点突变的方法,将An LPMO15g中3个N-糖基化修饰位点(151、334、385)上的天冬酰胺用中性谷氨酰胺取代。结果表明,作用于微晶纤维素时,突变体N-151产生的还原糖量比AnLPMO15g提高了10.34%,突变体N-385产生的还原糖量降低了12.73%,突变体N-334变化不显著;作用于稻草粉时,3个突变体产生的还原糖量均高于An LPMO15g,提高幅度为7%~19%。当与纤维素酶共同作用于微晶纤维素时,只有突变体N-334产生的还原糖量较AnLPMO15g显著提高了38.89%;共同作用于稻草粉时,突变体N-334和N-385产生的还原糖量略高于AnLPMO15g。由此可见,N-糖基化修饰对AnLPMO15g及其与纤维素酶协同降解作用均有不同程度的影响,其中N-151和N-385位点的糖基化修饰对提高An LPMO15g与纤维素酶协同降解活性尤为重要。
Auxiliary active family 9(AA9) in lytic polysaccharide monooxygenases(LPMOs) can effectively promote lignocellulose degradation. In order to study the effect of N-glycosylation on the synergy of AA9 protein AnLPMO15g from Aspergillus niger and cellulase, site-directed mutagenesis was used to replace asparagine on three N-glycosylation sites(151, 334, 385) in AnLPMO15g with neutral glutamine. It was found that when the mutants acted alone on microcrystalline cellulose, the yield of reducing sugar produced by mutant N-151 was 10.34% higher than that by AnLPMO15g, while the amount of reducing sugar produced by mutant N-385 was decreased by 12.73% and that by mutant N-334 was not significant. The yields of reducing sugar produced by three mutants were all higher than that by AnLPMO15g with an increase of 7%~19% when acting on straw powder. When acting on microcrystalline cellulose together with cellulase, only the mutant N-334 produced a significant increase in the amount of reducing sugar compared to AnLPMO15g by 38.89%. The mutant N-334 and N-385 produced slightly more reducing sugar than AnLPMO15g when co-acting on straw powder. N-glycosylation modification shows different effects on AnLPMO15g and its synergism with cellulase in cellulose degradation, and the glycosylation modification of N-334 site is especially important for improving the synergistic degradation activity of AnLPMO15g and cellulase.
作者
马立娟
佟文哲
杜丽平
崔馨予
马清
肖冬光
MA Li-juan;TONG Wen-zhe;DU Li-ping;CUI Xin-yu;MA Qing;XIAO Dong-guang(College of Biotechnology, Tianjin University of Science & Technology, Tianjin 300457, China;Key Laboratory of Industrial Fermentation Microbiology Ministry of Education, Tianjin 300457, China)
出处
《高校化学工程学报》
EI
CAS
CSCD
北大核心
2019年第5期1148-1155,共8页
Journal of Chemical Engineering of Chinese Universities
基金
国家自然科学基金(31770625)
天津市自然科学基金重点项目(16JCZDJC31880)
关键词
N-糖基化
纤维素酶
定点突变
协同性
N-glycosylation
cellulase
site-directed mutagenesis
synergy
