BINDING OF DIHYDROSTREPTOMYCIN TO ESCHERICHIA-COLI RIBOSOMES - CHARACTERISTICS AND EQUILIBRIUM OF REACTION
BINDING OF DIHYDROSTREPTOMYCIN TO ESCHERICHIA-COLI RIBOSOMES - CHARACTERISTICS AND EQUILIBRIUM OF REACTION
复制标题
DOI：
10.1128/aac.2.4.294
复制
发表时间：
1972-01-01
影响因子：
4.9
通讯作者：
FLAKS, JG
中科院分区：
文献类型：
--
作者：
CHANG, FN;FLAKS, JG
本文研究了双氢链霉素与几种不同大肠杆菌核糖体及核糖体亚基的结合，测定了Mg ~（2+）和pH的依赖性，以及盐和多核苷酸的影响。与敏感菌株的核糖体和亚基结合的唯一要求是10 mmMg 2+。一价盐削弱了结合的方式类似于对核糖核酸的二级结构的影响，这是拮抗在一定程度上增加量的Mg 2+。结合的双氢链霉素可以很容易地被链霉素和任何具有药理活性的衍生物所交换，但不能被抗生素的片段或任何其他氨基糖苷类所交换。与链霉素敏感菌株的天然（流出）70 S核糖体的结合是快速的，在0 - 37 ℃范围内与温度相对无关。多核苷酸不刺激结合。当二氢链霉素浓度达到10− 5 m时，超过95%的天然70 S核糖体恰好与1个抗生素分子紧密结合，结合复合物在25 ℃时的K diss为9.4 × 10− 8 m。在25 ℃下，与70 S核糖体结合的热力学参数如下：ΔG° = −9.6 kcal/mole，ΔH° = −6.2 kcal/mole，ΔS° = +11.4熵单位/mole。发现K-12菌株的核糖体与其它菌株的核糖体对抗生素的亲和力存在差异。大肠杆菌链霉素抗性或依赖菌株的70 S核糖体或亚基以及敏感菌株的50 S亚基的结合不显著。与来自敏感菌株的30 S亚基的结合比与70 S颗粒的结合弱一个数量级，aKdissat 25 C为10− 6 m。聚尿苷酸刺激这种结合轻微，但不影响结合分子的亲和力。当抗生素浓度高于10− 5 m时，链霉素敏感的70和30 S颗粒会结合更多的抗生素分子，同时也会结合链霉素耐药和依赖链霉素的菌株的核糖体，以及所有菌株的50 S亚基，所有这些结合平衡的Kdiss都是10− 4 m。这种较弱的非特异性结合与涉及颗粒的聚集现象的开始一致，并且发生在与紧密结合抗生素的单个位点不同的位点。后一个位点在一步突变为高水平抗性或依赖性后完全丢失。
The binding of dihydrostreptomycin to ribosomes and ribosomal subunits of a number of differentEscherichia colistrains was studied, and the Mg2+andpH dependence, as well as the effect of salts and polynucleotides, was determined. The only requirement for binding with ribosomes and subunits from susceptible strains was 10 mmMg2+. Monovalent salts weakened the binding in a manner similar to the effects on ribonucleic acid secondary structure, and this was antagonized to some extent by increased amounts of Mg2+. Bound dihydrostreptomycin could be readily exchanged by streptomycin and any antibiotically active derivative, but not by fragments of the antibiotic or any other aminoglycoside. With native (run-off) 70Sribosomes from streptomycin-susceptible strains, the binding was rapid and relatively temperature independent over the range from 0 to 37 C. Polynucleotides did not stimulate the binding. With concentrations of dihydrostreptomycin up to 10−5m, greater than 95% of native 70Sribosomes bound exactly 1 molecule of the antibiotic tightly, with aKdissfor the bound complex at 25 C of 9.4 × 10−8m. The following thermodynamic parameters were found for the binding with 70Sribosomes at 25 C:ΔG° = −9.6 kcal/mole, ΔH° = −6.2 kcal/mole, and ΔS° = +11.4 entropy units/mole. Differences in affinity for the antibiotic were found between ribosomes of K-12 strains and those of otherE. colistrains. There was insignificant binding to 70Sribosomes or subunits from streptomycin-resistant or -dependent strains, and to 50Ssubunits from susceptible strains. The binding to 30Ssubunits from susceptible strains was weaker by an order of magnitude than that to the 70Sparticle, with aKdissat 25 C of 10−6m. Polyuridylic acid stimulated this binding slightly but did not influence the affinity of the bound molecule. At antibiotic concentrations above 10−5m, streptomycin-susceptible 70Sand 30Sparticles bound additional molecules of the antibiotic, and binding also occurred to ribosomes from streptomycin-resistant and -dependent strains, as well as to 50Ssubunits from all strains.Kdissfor all of these binding equilibria were [Formula: see text] 10−4m. This weaker non-specific binding coincided with the beginning of aggregation phenomena involving the particles, and occurred at sites distinct from the single site which binds the antibiotic tightly. This latter site was completely lost after the one-step mutation to high-level resistance or dependence.