Integrated Chemical BioPhysics Research, Faculty of Science, Universiti Putra Malaysia, Serdang, Selangor, Malaysia.
Nanomolecular Laboratory, Faculty of Science, Universiti Putra Malaysia, Serdang, Selangor, Malaysia.
PLoS One. 2020 Nov 4;15(11):e0238147. doi: 10.1371/journal.pone.0238147. eCollection 2020.
Laccases, oxidative copper-enzymes found in fungi and bacteria were used as the basis in the design of nona- and tetrapeptides. Laccases are known to be excellent catalysts for the degradation of phenolic xenobiotic waste. However, since solvent extraction of laccases is environmentally-unfriendly and yields obtained are low, they are less preferred compared to synthetic catalysts. The histidine rich peptides were designed based on the active site of laccase extracted from Trametes versicolor through RCSB Protein Data Bank, LOMETS and PyMol software. The peptides were synthesized using Fmoc-solid phase peptide synthesis (SPPS) with 30-40% yield. These peptides were purified and characterized using LC-MS (purities >75%), FTIR and NMR spectroscopy. Synthesized copper(II)-peptides were crystallized and then analyzed spectroscopically. Their structures were elucidated using 1D and 2D NMR. Standards (o,m,p-cresol, 2,4-dichlorophenol) catalysed using laccase from Trametes versicolor (0.66 U/mg) were screened under different temperatures and stirring rate conditions. After optimizing the degradation of the standards with the best reaction conditions reported herein, medications with phenolic and aromatic structures such as ibuprofen, paracetamol (acetaminophen), salbutamol, erythromycin and insulin were screened using laccase (positive control), apo-peptides and copper-peptides. Their activities evaluated using GC-MS, were compared with those of peptide and copper-peptide catalysts. The tetrapeptide was found to have the higher degradation activity towards salbutamol (96.8%) compared with laccase at 42.8%. Ibuprofen (35.1%), salbutamol (52.9%) and erythromycin (49.7%) were reported to have the highest degradation activities using Cu-tetrapeptide as catalyst when compared with the other medications. Consequently, o-cresol (84%) was oxidized by Tp-Cu while the apo-peptides failed to oxidize the cresols. Copper(II)-peptides were observed to have higher catalytic activity compared to their parent peptides and the enzyme laccase for xenobiotic degradation.
漆酶是一种存在于真菌和细菌中的氧化铜酶，被用作设计九肽和四肽的基础。已知漆酶是酚类异生物质废物降解的优良催化剂。然而，由于溶剂萃取漆酶对环境不友好，且收率较低，因此与合成催化剂相比，它们的应用较少。基于从彩绒革盖菌中提取的漆酶的活性位点，通过 RCSB 蛋白质数据银行、LOMETS 和 PyMol 软件设计了富含组氨酸的肽。使用 Fmoc-固相肽合成（SPPS）以 30-40%的产率合成这些肽。使用 LC-MS（纯度>75%）、FTIR 和 NMR 光谱对合成的铜（II）-肽进行纯化和表征。合成的铜（II）-肽被结晶，然后进行光谱分析。使用 1D 和 2D NMR 阐明了它们的结构。使用 Trametes versicolor 中的漆酶（0.66 U/mg）对标准品（邻、间、对甲酚、2,4-二氯苯酚）进行筛选，在不同的温度和搅拌速率条件下进行。在优化了本文报道的最佳反应条件下标准品的降解后，使用漆酶（阳性对照）、脱肽和铜肽筛选具有酚和芳族结构的药物，如布洛芬、对乙酰氨基酚（扑热息痛）、沙丁胺醇、红霉素和胰岛素。使用 GC-MS 评估它们的活性，并将其与肽和铜肽催化剂的活性进行比较。与漆酶相比，四肽对沙丁胺醇的降解活性更高（96.8%）。与其他药物相比，当使用 Cu-四肽作为催化剂时，布洛芬（35.1%）、沙丁胺醇（52.9%）和红霉素（49.7%）被报道具有最高的降解活性。因此，Tp-Cu 氧化了邻甲酚（84%），而脱肽未能氧化甲酚。与母体肽和酶漆酶相比，铜（II）-肽对异生物质的降解表现出更高的催化活性。
