Biodesulfurization using thermophilic bacteria and identification of bds genes

嗜热菌生物脱硫及bds基因鉴定

• 批准号: 13650859
• 负责人: KIRIMURA Kohtaro
• 金额: $ 2.3万
• 依托单位: Waseda University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13650859/
• 关键词: Biotechnology; Bioengineering; Bioprocess; Microbial function; Biodesulfurization; Thermophilic bacteria; Dibenzothiophene; Flavin reductase; 環境微生物; Mycobacterium; 有機硫黄化合物; biodesulfurization; Bacillus subtilis; bioconversion; dibenzothiophe

Recalcitrant organosulfur compounds such as dibenzothiophene (DBT) derivatives in light gas oil (LGO) cannot be removed by hydrodesulfurization using metallic catalysts. The thermophilic bacteria, Bacillus subtilis WU-S2B and Mycobacterium phlei WU-F1, were isolated for its ability to grow on DBT as the sole source of sulfur at 50℃. In addition to DBT, WU-S2B and WU-F1 also could desulfurize alkylated DBTs such as 4, 6-dimethyl DBT through a sulfur-specific degradation pathway with the selective cleavage of carbon-sulfur bonds. Moreover, when resting cells of WU-F1 were incubated at 45℃ with two types of hydrodesulfurized LGOs in the reaction mixtures containing 50%(v/v)oils, the biodesulfurization reduced the sulfur contents from 120 to 50 ppm S (F-LGO) and from 34 to 15 ppm S (X-LGO), respectively. Gas chromatography analysis with an atomic emission detector revealed that the peaks of alkylated DBTs including 4-methyl DBT, 4, 6-dimethyl DBT, and 3, 4, 6-trimethyl DBT significantly decreased after the biodesulfurization. The DBT-desulfurization genes, bdsABC, were cloned from these two bacteria, and the flavin reductase genes, frb and frm, were also cloned from WU-S2B and Wu-F1, respectively. The recombinant WU-F1 carrying one more set of bdsABC and frm was constructed, and the desulfurizing activity of the recombinant strain was 2. 2-fold higher than that of the wild type strain WU-F1.

轻气油（LGO）中的顽固性有机硫化合物如二苯并噻吩（DBT）衍生物不能通过金属催化剂加氢脱硫去除。在50℃条件下，嗜热细菌枯草芽孢杆菌WU-S2B和真菌分枝杆菌WU-F1能够在单硫源DBT上生长。除DBT外，WU-S2B和WU-F1还可以通过选择性裂解碳硫键的硫特异性降解途径脱硫烷基化DBT，如4,6 -二甲基DBT。另外，在含50%（v/v）油的反应混合物中，将WU-F1的静息细胞与两种加氢脱硫的lgo在45℃下培养，生物脱硫的硫含量分别从120 ppm S （F-LGO）降至50 ppm S (X-LGO)，从34 ppm S降至15 ppm S （X-LGO）。原子发射检测器气相色谱分析表明，4-甲基DBT、4,6 -二甲基DBT和3,4,6 -三甲基DBT的烷基化峰在生物脱硫后显著降低。从这两种细菌中克隆出了dbt脱硫基因bdsABC，从WU-S2B和Wu-F1中分别克隆出了黄素还原酶基因frb和from。重组菌株WU-F1多携带一组bdsABC和frm，重组菌株的脱硫活性为2。比野生型菌株WU-F1高2倍。

项目成果

Kohtaro Kirimura et al.: "Identification and functional analysis of the genes encoding dibenzothiophene-desulfurizing enzymes from thermophilic bacteria"Appl.Microbiol.Biotechnol.. (In press). (2004)

Kohtaro Kirimura 等人：“来自嗜热细菌的编码二苯并噻吩脱硫酶的基因的鉴定和功能分析”Appl.Microbiol.Biotechnol..（正在出版）。

Kohtaro Kirimura, et al.: "Thermophilic biodesulfurization of hydrodesulfurized light gas oils by Mycobacterium Phlei WU-F1"FEMS Microbiol.Lett.. 221. 137-142 (2003)

Kohtaro Kirimura 等：“通过 Mycobacter Phlei WU-F1 对加氢脱硫轻质瓦斯油进行嗜热生物脱硫”FEMS Microbiol.Lett.. 221. 137-142 (2003)

Kohtaro Kirimura, et al.: "Thermophilic biosedulfurization of naphthothiophene and 2-ethylnaphthothiophene by a dibenzothiophene-desulfurizing bacterium, Mycobacterium phlei WU-F1"Appl.Microbiol.Biotechnol.. 58. 237-240 (2002)

Kohtaro Kirimura 等人：“二苯并噻吩脱硫细菌，草分枝杆菌 WU-F1 对萘并噻吩和 2-乙基萘并噻吩的高温生物硫化”Appl.Microbiol.Biotechnol.. 58. 237-240 (2002)

Kohtaro Kirimura, et al.: "Thermophilic biodesulfurization of dibenzothiophene and its derivatives by Mycobacterium phlei WU-F1"FEMS Microbiol.Lett.. 204. 129-133 (2001)

Kohtaro Kirimura 等：“草分枝杆菌 WU-F1 对二苯并噻吩及其衍生物的嗜热生物脱硫”FEMS Microbiol.Lett.. 204. 129-133 (2001)

Kohtaro Kirimura et al.: "Cloning of a gene encoding flavin reductase coupling with dibenzothiophene monooxygenase through coexpression screening using indigo production as selective indication"Biochem. Biophys. Res. Commun.. 313. 570-575 (2004)

Kohtaro Kirimura 等人：“通过使用靛蓝生产作为选择性指示的共表达筛选，克隆编码黄素还原酶与二苯并噻吩单加氧酶偶联的基因”Biochem。