QUENCHING OF THE INTRINSIC TRYPTOPHAN FLUORESCENCE OF MITOCHONDRIAL UBIQUINOL CYTOCHROME-C REDUCTASE BY THE BINDING OF UBIQUINONE
QUENCHING OF THE INTRINSIC TRYPTOPHAN FLUORESCENCE OF MITOCHONDRIAL UBIQUINOL CYTOCHROME-C REDUCTASE BY THE BINDING OF UBIQUINONE
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DOI：
10.1111/j.1432-1033.1988.tb13761.x
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发表时间：
1988-01-15
期刊：
影响因子：
--
通讯作者：
LENAZ, G
中科院分区：
文献类型：
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作者：
SAMWORTH, CM;DEGLIESPOSTI, M;LENAZ, G
1.泛醌(Q)对泛喹酚-细胞色素-c还原酶(复合体III)中色氨酸残基的本征荧光的猝灭已被用来提供关于线粒体呼吸链的这两个组成部分之间相互作用的直接信息。2.对荧光猝灭数据进行了内滤波校正，并用经典的Stern-Volmer图和修正的Stern-Volmer图进行了解释。这些曲线图的后者允许计算二联苯二酚和络合物III之间形成络合物的解离常数(Kd)，以及可被猝灭的荧光团的百分比。3.研究发现，不同的Q同系物以不同的亲和力结合到络合物III上，这取决于类异戊二烯链的长度：2，3-dimethoxy-5-methyl-6-decyl-1，4-benzoquinone，是O2的类似物，其Kd与Q2相同。此外，Q1中荧光团对猝灭的可及性低于所测试的其他对苯二酚。4.Q2与络合物III的结合亲和力取决于酶的氧化还原状态。5.加入络合物III抑制剂安替米星对荧光团的结合亲和力或猝灭剂的可及性影响很小。6.加入抗坏血酸抑制剂霉唑具有类似的还原抗坏血酸复合体III的效果。7.在氧化还原状态和抑制剂的添加方面，将络合物III重组为阿索莱菌素脂囊，得到了与溶液中的酶类似的定性结果。
1. The quenching by ubiquinone (Q) of the intrinsic fluorescence of tryptophan residues within ubiquinol-cytochrome-c reductase (complex III) has been exploited to provide direct information on the interaction between these two components of the mitochondrial respiratory chain. 2. The fluorescence quenching data have been corrected for inner filter effects and interpreted using the classical Stern-Volmer and modified Stern-Volmer plots. The latter of these plots allows computation of both the dissociation constant (Kd) of complex formation between ibiquinone and complex III, and the percentage of fluorophores accessible to quenching. 3. It is found that different Q homologues bind to complex III with different affinities depending upon the length of the isoprenoid chain: 2,3-dimethoxy-5-methyl-6-decyl-1,4-benzoquinone, an analogue of O2, exhibits the same Kd as Q2. Furthermore, the accessibility of fluorophores to quenching was lower for Q1 than for the other quinones tested. 4. The binding affinity of Q2 to complex III depends upon the redox state of the enzyme. 5. Addition of the complex III inhibitor, antimycin, has very little effect on the binding affinity or on the accessibility of fluorophores to the quencher. 6. Addition of the inhibitor myxothiazol has a similar effect of reducing complex III with ascorbate. 7. Reconstitution of complex III into asolectin lipid vesicles given similar qualitative results to the enzyme in solution regarding both the redox state and the addition of inhibitors.