Moura Amanda M A, Oliveira Jose Tadeu A, Sousa Daniele O B, Dias Lucas P, Araujo Nadine M S, de O Rocha Raquel, Aguiar Tawanny K B, Neto Joao M M, Silva Viviane O, Feitosa Ricardo M, Chaves Queilane L S G, Ramos Marcio V, Freitas Cleverson D T
Department of Biochemistry and Molecular Biology, Federal University of Ceará, Campus do Pici, Fortaleza-Ceará, Brazil. CEP 60451-970.
Department of Plant Pathology and Ecology, The Connecticut Agricultural Experiment Station, New Haven-CT 06511, USA.
Curr Protein Pept Sci. 2025;26(4):308-319. doi: 10.2174/0113892037339021241017084509.
Clinic infections caused by various microorganisms are a public health concern. The rise of new strains resistant to traditional antibiotics has exacerbated the problem. Thus, the search for new antimicrobial molecules remains highly relevant.
The current study purified, characterized, and assessed the antimicrobial activity of a papain inhibitor from L. seeds.
The inhibitor was purified by heating the crude extract at 80°C for 30 min, followed by ion exchange chromatography on a DEAE cellulose column. The purification index was 9-fold, yielding 2.3%. SDS-PAGE and size exclusion chromatography revealed that the protease inhibitor (PI) is a 15.9 kDa monomeric protein. The inhibition kinetics showed that PI is a competitive inhibitor specific to papain (Ki = 1.02 x 10 M). PI remained active even after heating at 100oC for 120 min and at pH conditions varying from 2.0 to 10.0. Even after 60 min, PI was resistant to papain proteolysis. PI exhibited antimicrobial activity against and Conclusion: Here, we show that PI is a highly stable type-1 cystatin with the potential to combat infections caused by and . Additional investigations into PI's structural aspects and mechanism of action, as well as safety assessments, are essential prerequisites for its potential application as a novel therapeutic intervention.
由各种微生物引起的临床感染是一个公共卫生问题。对传统抗生素耐药的新菌株的出现加剧了这一问题。因此，寻找新的抗菌分子仍然高度相关。
本研究对来自L.种子的木瓜蛋白酶抑制剂进行了纯化、表征和抗菌活性评估。
通过将粗提物在80°C加热30分钟，然后在DEAE纤维素柱上进行离子交换色谱法来纯化抑制剂。纯化指数为9倍，产率为2.3%。SDS-PAGE和尺寸排阻色谱显示蛋白酶抑制剂（PI）是一种15.9 kDa的单体蛋白。抑制动力学表明PI是木瓜蛋白酶的竞争性抑制剂（Ki = 1.02 x 10 M）。即使在100°C加热120分钟以及在pH值从2.0到10.0变化的条件下，PI仍保持活性。即使经过60分钟，PI也对木瓜蛋白酶水解具有抗性。PI对 和 表现出抗菌活性。结论：在此，我们表明PI是一种高度稳定的1型半胱氨酸蛋白酶抑制剂，具有对抗由 和 引起的感染的潜力。对PI的结构方面和作用机制进行进一步研究以及安全性评估，是其作为新型治疗干预措施潜在应用的必要前提条件。
