Miyamoto Y, Tiruppathi C, Ganapathy V, Leibach F H
Department of Cell and Molecular Biology, Medical College of Georgia, Augusta 30912-2100.
Biochim Biophys Acta. 1989 Jan 16;978(1):25-31. doi: 10.1016/0005-2736(89)90493-8.
The role of thiol groups in H+-gradient-dependent dipeptide transport in rabbit renal brush-border membrane vesicles was investigated using glycylsarcosine as the substrate. Treatment of the membrane vesicles with a thiol-group-reducing agent, dimercaptopropanol, stimulated Gly-Sar transport. On the other hand, treatment with thiol group oxidants such as 5,5'-dithiobis(2-nitrobenzoic acid), plumbagin and phenazine methosulfate inhibited Gly-Sar transport. These effects were irreversible, because washing the membranes after treatment failed to reverse the effects. Incubation of the membrane vesicles with phenylarsine oxide, a reagent which interacts specifically with vicinal dithiols, significantly inhibited Gly-Sar transport. In all cases, the stimulation or the inhibition of the dipeptide transport was primarily due to changes in the maximal velocity of the transport system, the apparent affinity constant remaining unaltered. These results demonstrate the involvement of one or more vicinal dithiol groups in the function of the renal dipeptide transport system and that these thiol groups must exist in reduced form to maintain maximal transport activity. In addition, these data indirectly suggest that a dithiol-disulfide interchange may play a role in the function of the renal dipeptide transport system.
以甘氨酰肌氨酸为底物，研究了巯基在兔肾刷状缘膜囊泡中H⁺梯度依赖性二肽转运中的作用。用巯基还原剂二巯基丙醇处理膜囊泡，可刺激甘氨酰肌氨酸转运。另一方面，用巯基氧化剂如5,5'-二硫代双(2-硝基苯甲酸)、白花丹素和吩嗪硫酸甲酯处理则抑制甘氨酰肌氨酸转运。这些作用是不可逆的，因为处理后洗涤膜未能逆转这些作用。用苯胂氧化物（一种与邻二硫醇特异性相互作用的试剂）孵育膜囊泡，可显著抑制甘氨酰肌氨酸转运。在所有情况下，二肽转运的刺激或抑制主要是由于转运系统最大速度的变化，表观亲和常数保持不变。这些结果表明一个或多个邻二硫醇基团参与了肾二肽转运系统的功能，并且这些巯基必须以还原形式存在以维持最大转运活性。此外，这些数据间接表明二硫醇-二硫化物交换可能在肾二肽转运系统的功能中起作用。
