Ubeid Anan Abu, Do Sylvia, Nye Chris, Hantash Basil M
Elixir Institute of Regenerative Medicine, 5941 Optical Court, San Jose, CA 95138, USA.
Biochim Biophys Acta. 2012 Oct;1820(10):1481-9. doi: 10.1016/j.bbagen.2012.05.003. Epub 2012 May 17.
Abnormal production and accumulation of melanin are characteristics of a number of skin disorders, including postinflammatory hyperpigmentation and melasma. Our objective was to develop and validate novel oligopeptides with potent inhibitory activity against mushroom and human tyrosinase with minimal toxicity toward melanocytes, keratinocytes, and fibroblasts.
A library of short sequence oligopeptides was docked against the crystal structure of mushroom tyrosinase to screen for favorable binding free energies and direct interaction with the catalytic pocket. The inhibitory activity of the octapeptides and hydroquinone (HQ) was assessed using mushroom and human tyrosinase and melanin content via human primary melanocytes. Effects on cell viability and proliferation were determined using the MTT assay and cytotoxicity via trypan blue exclusion.
Octapeptides P16-18 outperformed HQ, the benchmark of hypopigmenting agents, in all tested categories. Prolonged incubation of human keratinocytes, fibroblasts, or melanocytes with 30-3000μM HQ led to 8- to 65-fold greater cell death than with octapeptides. After 6d of incubation with 30μM HQ, we observed 70±3% and 60±2% cell death in melanocytes and fibroblasts, respectively, versus minimal toxicity up to an octapeptide concentration of 3mM.
Octapeptides P16-18 are potent competitive tyrosinase inhibitors with minimal toxicity toward the major cell types of human skin.
The findings in our study suggest that all three novel octapeptides may serve as safe and efficacious replacements of HQ for the treatment of pigmentary disorders.
黑色素的异常生成和积累是多种皮肤疾病的特征，包括炎症后色素沉着和黄褐斑。我们的目标是开发并验证具有强大抑制活性的新型寡肽，该寡肽对蘑菇酪氨酸酶和人酪氨酸酶具有抑制作用，同时对黑素细胞、角质形成细胞和成纤维细胞的毒性最小。
将短序列寡肽文库与蘑菇酪氨酸酶的晶体结构进行对接，以筛选出有利的结合自由能以及与催化口袋的直接相互作用。使用蘑菇酪氨酸酶和人酪氨酸酶以及通过人原代黑素细胞检测黑色素含量，评估八肽和对苯二酚（HQ）的抑制活性。使用MTT法测定对细胞活力和增殖的影响，并通过台盼蓝排斥法测定细胞毒性。
在所有测试类别中，八肽P16 - 18的表现均优于色素沉着抑制剂的基准对苯二酚。将人角质形成细胞、成纤维细胞或黑素细胞与30 - 3000μM的对苯二酚长时间孵育导致的细胞死亡比与八肽孵育时高8至65倍。用30μM对苯二酚孵育6天后，我们观察到黑素细胞和成纤维细胞的细胞死亡率分别为70±3%和60±2%，而在八肽浓度高达3mM时毒性最小。
八肽P16 - 18是有效的竞争性酪氨酸酶抑制剂，对人类皮肤的主要细胞类型毒性最小。
我们研究中的发现表明，所有三种新型八肽可能作为对苯二酚的安全有效替代品用于治疗色素沉着紊乱。
