论文部分内容阅读
在力复霉素ＳＶ研究中，发现硝酸盐对抗生素合成呈现多效性作用，不仅大幅度提高产量，还对产生菌——地中海拟无枝菌酸菌生理产生多方面的影响，从而提出整体性调节的结论．这一多效性作用是由硝酸盐所引起的，为此对硝酸还原酶进行了研究．首先，通过原生质体渗透裂解发现地中海拟无枝菌酸菌Ｕ－３２的硝酸还原酶是一个胞质酶．该酶极不稳定，缓冲液中加入硝酸钾、甘油等保护剂能极大地提高其稳定性．通过硫酸鱼精蛋白沉淀，硫酸铵分级分离，Ｐｈｅｎｙｌ－ＳｅｐｈａｒｏｓｅＣＬ４Ｂ、Ｂｉｏ－ＧｅｌＡ１．５ｍＤＥＡＥ－Ｓｅｐｈａｃｅｌ和ＳｅｐｈａｄｅｘＧ－７５柱层析等多步纯化得到了电泳纯的硝酸还原酶．该酶为一７９ｋＤ的单亚基酶，每分子酶含有约２．２９原子的钼，但并不含有非血红素铁、酸不稳定硫、ＦＭＮ及ＦＡＤ，其等电点为６．２，反应最适ｐＨ为７．２，最适温度为４０℃．对硝酸根的Ｋｍ值为１３．３μｍｏｌ／Ｌ．同时分析了该酶的吸收光谱．
In the casein regulins SV study, nitrite was found to have a pleiotropic effect on antibiotic synthesis, which not only greatly increased the yield, but also produced various physiological effects on the fungus-Mediterranean Amycolatopsis spp Conclusion of sexual adjustment. This pleiotropic effect is caused by nitrate, for which nitrate reductase has been studied. First, nitrate reductase, a member of U. mediterranei U-32, was found to be a cytosolic enzyme by protoplast permeation lysis. The enzyme is very unstable, adding buffer potassium nitrate, glycerol and other protective agents can greatly improve its stability. Electrophoretic pure nitrate reductase was obtained by multiple steps of protamine sulfate precipitation, ammonium sulfate fractionation, Phenyl-Sepharose CL4B, Bio-GelA1.5mDEAE-Sephacel and Sephadex G-75 column chromatography. The enzyme is a 79 kD mono-subunit enzyme containing about 2.29 atoms of molybdenum per molecule but does not contain non-heme iron, acid-labile sulfur, FMN and FAD with an isoelectric point of 6.2, The optimum reaction pH was 7.2, the optimum temperature was 40 ℃. The Km value for nitrate was 13.3 μmol / L. At the same time, the absorption spectrum of the enzyme was analyzed.