Abstract：
Ceramidases (CDases) are important in controlling skin barrier integrity by regulating ceramide composition and affording downstream signal molecules. While the functions of epidermal CDases are known, roles of neutral CDases secreted by skin-residing microbes are undefined. Here, we developed a one-step fluorogenic substrate, S-B, for specific detection of bacterial CDase activity and inhibitor screening. We identified a non-hydrolyzable substrate mimic, C6, as the best hit. Based on C6, we designed a photoaffinity probe, JX-1, which efficiently detects bacterial CDases. Using JX-1, we identified endogenous low-abundance PaCDase in a P. aeruginosa monoculture and in a mixed skin bacteria culture. Harnessing both S-B and JX-1, we found that CDase activity positively correlates with the relative abundance of P. aeruginosa and is negatively associated with wound area reduction in clinical diabetic foot ulcer patient samples. Overall, our study demonstrates that bacterial CDases are important regulators of skin ceramides and potentially play a role in wound healing.
摘要：
神经酰胺酶(CDases)在通过调节神经酰胺组成和提供下游信号分子来控制皮肤屏障完整性方面是重要的。虽然表皮CDases的功能是已知的，由皮肤微生物分泌的中性CDases的作用尚不明确。这里,我们开发了一步荧光底物，S-B,用于特异性检测细菌CDase活性和抑制剂筛选。我们鉴定了不可水解的底物模拟物C6作为最佳命中。基于C6，我们设计了一种光亲和探针JX-1，并观察到细菌CDases的有效检测。使用JX-1，我们在铜绿假单胞菌单一培养和混合皮肤细菌培养物中鉴定了内源性低丰度PaCDase。利用S-B和JX-1,我们在临床糖尿病足溃疡患者样品中观察到CDase活性与铜绿假单胞菌相对丰度正相关,与伤口面积减少负相关。总的来说,我们的研究表明,细菌CDases是皮肤神经酰胺的重要调节因子,可能在伤口愈合中发挥作用.
