Wang Yuguang, Liu Qian, Zhou Hongbo, Chen Xinhua
Key Laboratory of Marine Biogenetic Resources, Third Institute of Oceanography, State Oceanic Administration, Xiamen, 361005 People's Republic of China.
Fujian Collaborative Innovation Center for Exploitation and Utilization of Marine Biological Resources, Xiamen, 361005 People's Republic of China.
3 Biotech. 2017 Dec;7(6):360. doi: 10.1007/s13205-017-0995-z. Epub 2017 Oct 3.
The cysteine desulfurase (SufS) gene of TPY, a Gram-positive bacterium isolated from deep-sea hydrothermal vent, was cloned and over-expressed in BL21. The recombinant SufS protein was purified by one-step affinity chromatography. The TPY SufS contained a well conserved motif RXGHHCA as found in that of other microorganisms, suggesting that it belonged to group II of cysteine desulfurase family. The recombinant TPY SufS could catalyze the conversion of l-cysteine to l-alanine and produce persulfide, and the enzyme activity was 95 μ/μL of sulfur ion per minute. The growth of BL21 was promoted by over-expressing TPY SufS in vivo or by directly adding recombinant TPY SufS in the medium (4.3-4.5 × 10 cells/mL vs. 3.2-3.5 × 10 cells/mL). Furthermore, the highest cell density of BL21 when the TPY SufS was over-expressed was about 3.5 times that of the control groups in the presence of sodium thiosulfate. These results indicate that the SUF system as the only assembly system of iron-sulfur clusters not only has significant roles in survival of TPY, but also might be important for combating with high content of sulfide.
从深海热液喷口分离出的革兰氏阳性细菌TPY的半胱氨酸脱硫酶（SufS）基因被克隆并在BL21中过表达。重组SufS蛋白通过一步亲和层析法纯化。TPY SufS含有一个与其他微生物中发现的保守基序RXGHHCA，表明它属于半胱氨酸脱硫酶家族的第二组。重组TPY SufS可以催化L-半胱氨酸转化为L-丙氨酸并产生过硫化物，酶活性为每分钟95μ/μL硫离子。通过在体内过表达TPY SufS或通过在培养基中直接添加重组TPY SufS促进了BL21的生长（4.3 - 4.5×10细胞/mL对3.2 - 3.5×10细胞/mL）。此外，在存在硫代硫酸钠的情况下，过表达TPY SufS时BL21的最高细胞密度约为对照组的3.5倍。这些结果表明，作为铁硫簇唯一组装系统的SUF系统不仅在TPY的存活中具有重要作用，而且可能对于抵抗高含量的硫化物也很重要。
