Rationale-Based, De Novo Design of Dehydrophenylalanine-Containing Antibiotic Peptides and Systematic Modification in Sequence for Enhanced Potency

Abstract

Increased microbial drug resistance has generated a global requirement for new anti-infective agents. As part of an effort to develop new, low-molecular-mass peptide antibiotics, we used a rationale-based minimalist approach to design short, nonhemolytic, potent, and broad-spectrum antibiotic peptides with increased serum stability. These peptides were designed to attain an amphipathic structure in helical conformations. VS1 was used as the lead compound, and its properties were compared with three series of derivates obtained by (i) N-terminal amino acid addition, (ii) systematic Trp substitution, and (iii) peptide dendrimerization. The Trp substitution approach underlined the optimized sequence of VS2 in terms of potency, faster membrane permeation, and cost-effectiveness. VS2 (a variant of VS1 with two Trp substitutions) was found to exhibit good antimicrobial activity against both the Gram-negative Escherichia coli and the Gram-positive bacterium Staphylococcus aureus. It was also found to have noncytolytic activity and the ability to permeate and depolarize the bacterial membrane. Lysis of the bacterial cell wall and inner membrane by the peptide was confirmed by transmission electron microscopy. A combination of small size, the presence of unnatural amino acids, high antimicrobial activity, insignificant hemolysis, and proteolytic resistance provides fundamental information for the de novo design of an antimicrobial peptide useful for the management of infectious disease.

Rationally Designed Transmembrane Peptide Mimics of the Multidrug Transporter Protein Cdr1 Act as Antagonists to Selectively Block Drug Efflux and Chemosensitize Azole-resistant Clinical Isolates of Candida albicans

“…TMPM Synthesis and Purification-Synthesis and purification of the TMPMs and FITC-labeled TMPMs were performed as described previously (10). Briefly, TMPMs were synthesized (1 mM synthesis scale) using solid-phase methods with Fmoc chemistry on Rink amide-MBHA (4-methylbenzhydrylamine hydrochloride salt) resin in the manual mode using N,N-diisopropylcarbodiimide and N-hydroxybenzotriazole as coupling reagents.…”

Plasmodium falciparum Merozoite Surface Protein 3

“…Peptide Synthesis-The pentapeptide sequence from MSP3 protein (YILGW) determined as oligomerization motif by TANGO algorithm was synthesized by the methods described previously (23). Peptide was synthesized on Rink-amide 4-methylbenzhydrylamine resin (0.75 mmol/g) using Fmoc methodology.…”

In vitro antibacterial and antimalarial activity of dehydrophenylalanine-containing undecapeptides alone and in combination with drugs

“…Rifampicin, a non-peptide antibiotic, has a MIC of 1 M against S. aureus [10]. An additive effect was also observed against S. aureus when VA1 and VA3 were used in combination with 500 nM rifampicin (MIC 50 ).…”

“…Both the coupling of amino acids and the Fmoc deprotection were monitored by the Kaiser test [13]. The amino acid dehydrophenylalanine ( Phe) was introduced into peptides as an Fmoc-X-Phe azalactone [where X is Lys (Boc), Trp (Boc) or Ala] dipeptide block, which was allowed to couple overnight in DMF [10,12].…”

“…Synthetic or designer AMPs have recently become a focus for drug design owing to high-throughput studies that generated multiple synthetic peptides active against a broad range of pathogens [8,9]. In our previous work, we have reported the de novo design of a series of AMPs with activity towards the Gram-negative bacterium Escherichia coli and the Gram-positive bacterium Staphylococcus aureus [10]. These peptides incorporated a non-natural amino acid, dehydrophenylalanine, which is the dehydro analogue of the amino acid phenylalanine and is found in the antibiotic peptides of fungal origin such as tentoxin [11].…”