新型抗 MRSA 肽的纯化与鉴定，由 sp. SPR-20 产生。

Purification and Characterization of Novel Anti-MRSA Peptides Produced by sp. SPR-20.

机构信息

School of Pharmacy, Walailak University, Nakhon Si Thammarat 80161, Thailand.

School of Allied Health Sciences, Walailak University, Nakhon Si Thammarat 80161, Thailand.

出版信息

Molecules. 2022 Dec 2;27(23):8452. doi: 10.3390/molecules27238452.

Methicillin-resistant (MRSA) is listed as a high-priority pathogen because its infection is associated with a high mortality rate. It is urgent to search for new agents to treat such an infection. Our previous study isolated a soil bacterium ( sp. SPR-20), showing the highest antimicrobial activity against TISTR 517 and MRSA strains. The present study aimed to purify and characterize anti-MRSA substances produced by SPR-20. The result showed that five active substances (P1-P5) were found, and they were identified by LC-MS/MS that provided the peptide sequences of 14-15 residues. Circular dichroism showed that all peptides contained β-strand and disordered conformations as the major secondary structures. Only P1-P4 adopted more α-helix conformations when incubated with 50 mM SDS. These anti-MRSA peptides could inhibit and MRSA in concentrations of 2-32 μg/mL. P1 (NH-VVVNVLVKVLPPPVV-COOH) had the highest activity and was identified as a novel antimicrobial peptide (AMP). The stability study revealed that P1 was stable in response to temperature, proteolytic enzymes, surfactant, and pH. The electron micrograph showed that P1 induced bacterial membrane damage when treated at 1× MIC in the first hour of incubation. The killing kinetics of P1 was dependent on concentration and time. Mechanisms of P1 on tested pathogens involved membrane permeability, leakage of genetic material, and cell lysis. The P1 peptide at a concentration up to 32 μg/mL showed hemolysis of less than 10%, supporting its safety for human erythrocytes. This study provides promising anti-MRSA peptides that might be developed for effective antibiotics in the post-antibiotic era.

耐甲氧西林金黄色葡萄球菌（MRSA）被列为高优先级病原体，因为其感染与高死亡率相关。因此，迫切需要寻找新的药物来治疗这种感染。我们之前的研究从土壤中分离出一株细菌（sp. SPR-20），该菌对 TISTR 517 和 MRSA 菌株表现出最高的抗菌活性。本研究旨在纯化和鉴定 SPR-20 产生的抗 MRSA 物质。结果表明，发现了五种活性物质（P1-P5），通过 LC-MS/MS 鉴定，提供了 14-15 个残基的肽序列。圆二色性表明，所有肽均含有β-折叠和无规卷曲构象，作为主要的二级结构。只有 P1-P4 在与 50mM SDS 孵育时采用更多的α-螺旋构象。这些抗 MRSA 肽可在 2-32μg/mL 的浓度下抑制 MSSA 和 MRSA。P1（NH-VVVNVLVKVLPPPVV-COOH）活性最高，被鉴定为一种新型抗菌肽（AMP）。稳定性研究表明，P1 对温度、蛋白酶、表面活性剂和 pH 均稳定。电子显微镜显示，当以 1×MIC 处理时，P1 在孵育的第一个小时内诱导细菌膜损伤。P1 的杀菌动力学取决于浓度和时间。P1 对测试病原体的作用机制包括膜通透性、遗传物质泄漏和细胞裂解。浓度高达 32μg/mL 的 P1 肽的溶血率小于 10%，支持其对人红细胞的安全性。本研究提供了有前景的抗 MRSA 肽，可能在抗生素后时代开发为有效的抗生素。