Guerrieri F, Papa S
J Bioenerg Biomembr. 1981 Dec;13(5-6):393-409. doi: 10.1007/BF00743212.
The effect of chemical modifiers of amino acid residues on the proton conductivity of H+-ATPase in "inside out" submitochondrial particles has been studied. Treatment of submitochondrial particles prepared in the presence of EDTA (ESMP) with the arginine modifiers, phenylglyoxal or butanedione, or the tyrosine modifier, tetranitromethane, caused inhibition of the ATPase activity. Phenylglyoxal and tetranitromethane also caused inhibition of the anaerobic release of respiratory delta micro H+ in ESMP as well as in particles deprived of F1 (USMP). Butanedione treatment caused, on the contrary, acceleration of anaerobic proton release in both particles. The inhibition of proton release caused by phenylglyoxal and tetranitromethane exhibition in USMP a sigmoidal titration curve. The same inhibitory pattern was observed with oligomycin and with N,N'-dicyclohexylcarbodiimide. In ESMP, relaxation of delta micro H+ exhibited two first-order phases, both an expression of the H+ conductivity of the ATPase complex. The rapid phase results from transient enhancement of H+ conduction caused by respiratory delta micro H+ itself. Oligomycin, N,N'-dicyclohexylcarbodiimide, and tetranitromethane inhibited both phases of H+ release, and butanedione accelerated both. Phenylglyoxal inhibited principally the slow phase of H+ conduction. In USMP, H+ release followed simple first-order kinetics. Oligomycin depressed H+ release, enhanced respiratory delta micro H+, and restored the biphasicity of H+ release. Phenylglyoxal and tetranitromethane inhibited H+ release in USMP without modifying its first-order kinetics. Butanedione treatment caused biphasicity of H+ release from USMP, introducing a very rapid phase of H+ release. Addition of soluble F1 to USMP also restored biphasicity of H+ release. A mechanism of proton conduction by Fo is discussed based on involvement of tyrosine or other hydroxyl residues, in series with the DCCD-reactive acid residue. There are apparently two functionally different species of arginine or other basic residues: those modified by phenylglyoxal, which facilitate H+ conduction, and those modified by butanedione, which retard H+ diffusion.
研究了氨基酸残基的化学修饰剂对“内翻”亚线粒体颗粒中H⁺ -ATP酶质子传导率的影响。用精氨酸修饰剂苯乙二醛或丁二酮，或酪氨酸修饰剂四硝基甲烷处理在乙二胺四乙酸（EDTA）存在下制备的亚线粒体颗粒（ESMP），会抑制ATP酶活性。苯乙二醛和四硝基甲烷也会抑制ESMP以及缺乏F1的颗粒（USMP）中呼吸性δ微H⁺的厌氧释放。相反，丁二酮处理会使两种颗粒中的厌氧质子释放加速。苯乙二醛和四硝基甲烷对质子释放的抑制在USMP中呈现出S形滴定曲线。用寡霉素和N，N'-二环己基碳二亚胺也观察到相同的抑制模式。在ESMP中，δ微H⁺的弛豫呈现出两个一级相，这两者都是ATP酶复合体H⁺传导率的表现。快速相是由呼吸性δ微H⁺本身引起的H⁺传导的短暂增强导致的。寡霉素、N，N'-二环己基碳二亚胺和四硝基甲烷抑制H⁺释放的两个相，而丁二酮则加速这两个相。苯乙二醛主要抑制H⁺传导的慢相。在USMP中，H⁺释放遵循简单的一级动力学。寡霉素抑制H⁺释放，增强呼吸性δ微H⁺，并恢复H⁺释放的双相性。苯乙二醛和四硝基甲烷在不改变其一级动力学的情况下抑制USMP中的H⁺释放。丁二酮处理使USMP中的H⁺释放呈现双相性，引入了一个非常快速的H⁺释放相。向USMP中添加可溶性F1也恢复了H⁺释放的双相性。基于酪氨酸或其他羟基残基与DCCD反应性酸性残基串联参与，讨论了F₀的质子传导机制。显然有两种功能不同的精氨酸或其他碱性残基：那些被苯乙二醛修饰的，促进H⁺传导；那些被丁二酮修饰的，阻碍H⁺扩散。
