Biodesulfurization using thermophilic bacteria and identification of bds genes

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。

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.