A New Synthetic Peptide with In vitro Antibacterial Potential Against Escherichia coli O157:H7 and Methicillin-Resistant Staphylococcus aureus (MRSA)

Abstract

In this work, we performed the rational design of a cationic antimicrobial peptide, GIBIMPY4, using the software DEPRAMPs developed at the GIBIM research group. GIBIMPY4 has a length of 17 amino acids, it is amphipathic, its structure is α-helix and it has a net charge of (+5). Solid-phase peptide synthesis was performed using the Fmoc strategy in acid medium. The primary structure was confirmed by MALDI-TOF mass spectrometry. The antimicrobial activity of the peptide was evaluated by broth microdilution method by measuring optical density in 96-well microplates. The minimal inhibitory concentration of GIBIMPY4 to kill 50 % of the bacterial cells (MIC50) was 6.20 ± 0.02 µM for MRSA and 4.55 ± 0.02 µM for E. coli O157:H7, while also reporting a bacteriostatic effect for the later. GIBIMPY4 activity was sensitive to salt concentration in E. coli but insignificant effect in its activity against MRSA. The peptide seems to be a broad-spectrum antimicrobial agent based on the results against Gram-positive and Gram-negative bacteria and was specific for bacterial cells E. coli O157:H7 with index of specificity equal to 9.01 in vitro assays.

Achatina fulicaHaemocyanin-Derived Peptides as Novel Antimicrobial Agents

“…Solid-phase chemical synthesis was performed employing the Fmoc strategy in polypropylene bags according to the methodology described by (Cruz et al, 2014;Guzmán et al, 2020;Prada et al, 2016). The peptides were lyophilized and characterized by high-performance liquid chromatography (HPLC) in a JASCO system (JASCO Corp., Tokyo, Japan), and molecular mass of was determined by electrospray-mass spectrometry (ESI-MS) in a LCMS-2020 ESI-MS equipment (Shimadzu Corp., Kyoto, Japan).…”

“…Haemolysis assay was conducted according to the method described by (Prada et al, 2016). Human erythrocytes were washed 4 times with PBS buffer and suspended at the ratio of 8x10 9 blood cells/mL.…”

How to Combat Gram-Negative Bacteria Using Antimicrobial Peptides: A Challenge or an Unattainable Goal?

“…The Universidad Industrial de Santander in Colombia has developed in silico tools such as DEPRAMPs software; this is based on a genetic algorithm and is currently being patented. Prada et al, used this algorithm to design the 17 residue-long GIBIMPY4 (SFIKRSLKLIKSLVLIK) peptide which is active against pathogenic bacteria such as E. coli O157:H7 and methicillin-resistant S. aureus , but has a considerable cytotoxic and haemolytic effect [ 102 ]. Other peptides having great antibacterial potential but low selectivity (haemolytic and cytotoxic effect) have been designed using this software [ 103 ], thereby highlighting the need to complement the design algorithm method with haemolytic and/or cytotoxic activity prediction for obtaining AMPs which are highly selective for bacterial cells.…”

Diseño, síntesis, caracterización y evaluación in vitro de la actividad de los péptidos antimicrobianos contra bacterias patógenas resistentes a antibióticos

“…Estructura, propiedades bioquímicas y actividad de los péptidos antimicrobianos. Aunque el número y la diversidad de los péptidos antimicrobianos en la naturaleza son elevadas (véase la base de datos en http://aps.unmc.edu/AP/main.php), hay formas estructurales que la mayoría de ellos comparten (Cruz, et al, 2014;2017;2018;Prada, et al, 2016;Kumar, et al, 2018). En general, los péptidos antimicrobianos son relativamente cortos (12 a 30 aminoácidos), de naturaleza catiónica (cargados positivamente con un valor de pH neutro) y anfifílicos, con una proporción significativa de residuos hidrofóbicos.…”

“…Se ha propuesto una variedad de estrategias para mejorar la estabilidad de los péptidos frente a la degradación proteolítica. Entre ellas, se han ensayado la modificación química de extremos terminales de péptidos (Danial, et al, 2012); la alteración de sus estructuras secundarias flexibles (Rozek, et al, 2003); la estabilización de las estructuras secundarias (Houston, et al, 1998); el desarrollo de péptidos análogos mediante la sustitución con aminoácidos no naturales (por ejemplo, usando el enantiómero D) (Lee, et al, 2011;Cruz, et al, 2018); el aumento del contenido alfa helicoidal en la estructura de péptidos mediante modificaciones covalentes (Uteng, et al, 2003); la modificación de la propiedad hidrófoba, la longitud y la carga neta (Rydlo, et al, 2006); el acortamiento de péptidos antimicrobianos naturales (Lee, et al, 2011), y el diseño in silico de péptidos antimicrobianos utilizando los algoritmos genéticos (Prada, et al, 2016). Además, han surgido estrategias de diseño racional basadas en el alineamiento de secuencias de los péptidos antimicrobianos en una plantilla.…”