离子和pH值对维吉尼亚霉素S与核糖体结合的作用。

Action of ions and pH on the binding of virginiamycin S to ribosomes.

作者信息

Di Giambattista M, Cocito C

出版信息

Biochim Biophys Acta. 1983 May 4;757(1):92-100. doi: 10.1016/0304-4165(83)90156-3.

Abstract

Virginiamycin S (VS) binds to the 50 S ribosomal subunits (KaVS = 2.5 X 10(6) M-1); the affinity of ribosomes for VS undergoes a 6-fold increase (KaVS = 15 X 10(6) M-1) in the presence of virginiamycin M (VM). In the present work, the action of inorganic ions and pH on the binding reaction of VS to ribosomes was analyzed by a spectrofluorimetric technique, in the presence and in the absence of VM. Preliminary to this study, the interaction of ions with free VS was also explored. In aqueous solution and in the absence of VM, chelation by VS of K+, NH4+, Na+, Mg2+ and Ca2+ was observed. The binding of Mg2+ was strongly influenced by monovalent ions and pH between 7 and 8: the association constants for the VS:Mg2+ complex were 12.5 M-1 (+NH4+) and 45 M-1 (-NH4+). Binding of VS to ribosomes occurred as well in the presence of NH4+ as with K+, but was suppressed upon replacement of these ions by Na+. The association constant of the VS binding reaction to ribosomes was strongly enhanced by VM in the presence of either NH4+ or K+ but, while a plateau value was observed over the entire range of NH4+ concentrations, 100-fold higher values were obtained at low (25 mM) K+. This VM-induced increase of ribosome affinity for VS (KaVS) was dependent also on Mg2+, which displayed a higher synergistic action with K+ than with NH4+ (the highest KaVS values were recorded for 5 mM Mg2+ and 100 mM K+). In this reaction, Mg2+ could be replaced by Ca2+, whereas spermidine was ineffective. Also, the VM-promoted enhancement of the KaVS value increased with the pH. In conclusion, the two ribosomal functions analyzed in the present work, VS-binding to ribosome and VM-promoted enhancement of ribosome affinity for VS, rely on the concentration of monovalent and bivalent ions and on the pH. Both functions require either NH4+ or K+, and either Mg2+ or Ca2+ (the elements of each couple are not truly equivalent), from which ribosome conformation depends.

摘要

维吉尼亚霉素S（VS）与50 S核糖体亚基结合（KaVS = 2.5×10⁶ M⁻¹）；在维吉尼亚霉素M（VM）存在的情况下，核糖体对VS的亲和力增加6倍（KaVS = 15×10⁶ M⁻¹）。在本研究中，采用荧光光谱技术分析了无机离子和pH值对VS与核糖体结合反应的影响，实验分别在有和没有VM的情况下进行。在这项研究之前，还探索了离子与游离VS的相互作用。在水溶液中且没有VM的情况下，观察到VS对K⁺、NH₄⁺、Na⁺、Mg²⁺和Ca²⁺的螯合作用。Mg²⁺的结合受到单价离子和7至8之间pH值的强烈影响：VS:Mg²⁺复合物的缔合常数分别为12.5 M⁻¹（+NH₄⁺）和45 M⁻¹（-NH₄⁺）。在NH₄⁺和K⁺存在的情况下，VS均能与核糖体结合，但被Na⁺取代这些离子后结合受到抑制。在NH₄⁺或K⁺存在的情况下，VM会强烈增强VS与核糖体结合反应的缔合常数，然而，在整个NH₄⁺浓度范围内观察到一个平稳值，而在低（25 mM）K⁺浓度下获得的值高出100倍。VM诱导的核糖体对VS亲和力（KaVS）的增加也依赖于Mg²⁺，Mg²⁺与K⁺的协同作用比与NH₄⁺的协同作用更强（在5 mM Mg²⁺和100 mM K⁺时记录到最高的KaVS值）。在这个反应中，Mg²⁺可以被Ca²⁺取代，而亚精胺无效。此外，VM促进的KaVS值的增加随着pH值的升高而增加。总之，本研究中分析的两种核糖体功能，即VS与核糖体的结合以及VM促进的核糖体对VS亲和力的增强，依赖于单价和二价离子的浓度以及pH值。这两种功能都需要NH₄⁺或K⁺，以及Mg²⁺或Ca²⁺（每对元素并非完全等效），核糖体的构象取决于这些元素。