球形红细菌降解HMX的途径及产酶特性  

Degradation Pathway of HMX and the Property of Crude Enzyme Produced by Rhodobacter sphaeroides

作　　者：白红娟 赵婷婷 康鹏洲 高莉 BAI Hong-juan;ZHAO Ting-ting;KANG Peng-zhou;GAO Li(School of Environment and Safety Engineering,North University of China,Taiyuan 030051,China)

机构地区：[1]中北大学环境与安全工程学院

出　　处：《含能材料》2019年第7期550-557,I0004,共9页Chinese Journal of Energetic Materials

基　　金：山西省回国留学人员科研资助项目(2016-084)

摘　　要：为了进一步研究球形红细菌(Rhodobacter sphaeroides)H菌株对HMX生物转化过程及产酶特性,研究了不同碳源、氮源及金属离子对该菌株生物转化奥克托今(HMX)的效率及其生长的影响;采用液相色谱-质谱联用仪(LC-MS)分析了球形红细菌降解HMX的中间代谢产物,推测了可能的降解途径;测定了不同条件对菌株产酶比活力的影响,用聚丙烯酰胺凝胶电泳法分析了酶谱。结果表明,该菌株转化HMX的最适碳源、组合氮源和金属离子分别为苹果酸、(NH4)2SO4和酵母膏以及Ca2+;当HMX的初始浓度为100mg·L^-1时,培养96h后,可以检测到三种物质:两种HMX亚硝基衍生物(单亚硝基mNs-HMX和二亚硝基dNs-HMX)及次甲基二硝胺(MEDINA),其母离子的质荷比分别为279、263和136。推测其可能的降解途径为两条,一条是利用还原酶,HMX被还原为mNs-HMX和dNs-HMX;另外一条,利用水解酶,HMX经转化并开环裂解为次甲基二硝胺。菌体细胞粗酶液酶比活力和聚丙烯酰胺凝胶电泳实验表明,HMX浓度在75,100mg·L^-1时,对菌株产酶比活力有明显的促进作用,而HMX浓度为125,150mg·L^-1时,对菌株产酶比活力有抑制作用。pH为7.0时,该菌株产生的酶比活力最高。To further study the biotransformation process and producing enzyme properties of bacterial strain of Rhodobacter sphaeroides to HMX,the effects of different carbon sources,nitrogen sources and metal ions on the biotransformation efficiency of HMX of the strain and its growth were studied. The intermediate metabolites of HMX degraded by Rhodobacter sphaeroides were analyzed by liquid chromatography-mass spectrometry(LC-MS),and the possible degradation pathways were presumed. The effects of different conditions on the specific activity of the enzymes produced by the strain were determined. The zymograms were analyzed by polyacrylamide gel electrophoresis. The results show that the optimum carbon source,combined nitrogen source and metal ion for transforming HMX by the strain are malic acid,(NH4)2SO4 and yeast extract,Ca2+,respectively. When the initial concentration of HMX is 100 mg·L^-1,after 96 h of culture,three substances can be detected:two HMX nitroso derivatives(mononitroso mNs-HMX and dinitroso dNs-HMX),methine dinitramine(MEDINA),and the mass-to-charge ratios of their mother ions are 279,263 and 136,respectively. The possible degradation pathways presumed have two branches. One branche is that HMX is reduced to mNs-HMX and dNs-HMX by reductase,the other branche is that HMX is transformed and open-loop cleavaged into methine dinitramine by hydrolase. The specific activity of enzyme and polyacrylamide gel electrophoresis experiments show that the specific activity of the enzyme produced by the strain is significantly promoted when the concen - tration of HMX is 75 mg·L^-1 and 100 mg·L^-1,while when the concentration of HMX is 125 and 50 mg·L^-1,the specific activity of the enzyme produced by the strain has an inhibitory effect. When pH is 7,the specific activity of the enzyme produced by the strain is the highest.

关 键 词：奥克托今(HMX) 球形红细菌 中间产物 降解途径 粗酶液分析 

分 类 号：TJ55[兵器科学与技术—军事化学与烟火技术] X172[环境科学与工程—环境科学]