INHIBITION OF YEAST RIBONUCLEIC-ACID POLYMERASES BY THIOLUTIN
INHIBITION OF YEAST RIBONUCLEIC-ACID POLYMERASES BY THIOLUTIN
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DOI：
10.1128/jb.116.1.245-256.1973
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发表时间：
1973-01-01
影响因子：
3.2
通讯作者：
TIPPER, DJ
中科院分区：
文献类型：
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作者：
TIPPER, DJ
酵母菌核糖核酸聚合酶II经二乙氨基乙基纤维素(DE-52)或DEAE-Sephadex(A-25)分离后，可被1.5μgα-Amanitin抑制50%。这种抑制作用与酶、模板、核苷酸和抗生素的相互作用顺序无关，当将α-Amanitin加入到活性合成核糖核酸的制剂中时，这种抑制立即表现出来。因此，α-Amanitin的主要作用是抑制这个系统中预先启动的RNAs序列的延长，就像在其他系统中一样。α-Amanitin抗性RNA聚合酶活性(I)的单峰在酶II之前在任一柱上洗脱。在A-25而不是在DE-52上，在酶II之后洗脱出第三个活性峰(III)。该活性也对α-Amanitin具有抗性。酶I、酶II和酶III分别被每毫升3、4和3μg硫代甲苯抑制50%。抑制的程度与模板的性质(天然或变性的鲑鱼精子脱氧核糖核酸或多聚(da-dT)或0.4 mM二硫苏糖醇的存在无关，但这种显著的抑制仅在没有模板的情况下与硫质素预孵育时才能看到。在没有核苷酸的情况下，模板保护酶不受硫蛋白的影响。要么在与模板相互作用之前，聚合酶上的敏感部位只被硫蛋白接触，要么硫蛋白抑制功能性聚合酶-模板相互作用，而不是预先启动的RNA链的延长。
Yeast ribonucleic acid (RNA) polymerase II, isolated after fractionation on diethylaminoethyl (DEAE)-cellulose (DE-52) or on DEAE-Sephadex (A-25), is 50% inhibited by 1.5 μg of α-amanitin. This inhibition is independent of the sequence of interaction of enzyme, template, nucleotides, and antibiotic and is expressed immediately on addition of α-amanitin to a preparation actively synthesizing RNA. Thus, α-amanitin's primary effect is inhibition of elongation of preinitiated RNA sequences in this system, as in others. A single peak of α-amanitin-resistant RNA polymerase activity (I) was eluted before enzyme II on either column. On A-25 but not on DE-52, a third peak of activity (III) was eluted after enzyme II. This activity was also resistant to α-amanitin. Enzymes I, II, and III were 50% inhibited by 3, 4, and 3 μg of thiolutin per ml, respectively. The extent of inhibition was independent of the nature of the template (native or denatured salmon sperm deoxyribonucleic acid or poly(dA-dT) or of the presence of 0.4 mM dithiothreitol, but this marked inhibition was only seen when enzymes were preincubated with thiolutin in the absence of template. Template protected the enzymes against thiolutin in the absence of nucleotides. Either the sensitive site on the polymerase is only accessible to thiolutin before interaction with template or thiolutin inhibits functional polymerase-template interaction but not elongation of preinitiated RNA chains.