Optimization of oncocin for antibacterial activity using a SPOT synthesis approach: extending the pathogen spectrum to Staphylococcus aureus
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The identification of lead molecules against multidrug-resistant bacteria ensuing the development of novel antimicrobial drugs is an urgent task. Proline-rich antimicrobial peptides are highly active in vitro and in vivo, but only against a few Gram-negative human pathogens, with rather weak activities against Pseudomonas aeruginosa and Staphylococcus aureus. This reduced level of efficacy could be related to inadequate uptake mechanisms or structural differences of the intracellular target proteins, i.e., the 70S ribosome or chaperone DnaK. Here we synthesized peptide arrays on cellulose membranes using cleavable linkers to release the free individual peptides for further antimicrobial tests. Thus, a library of singly substituted oncocin analogs was produced by replacing each residue by all other 19 canonical amino acids yielding a set of 361 individual peptides to be evaluated against a luminescent P. aeruginosa strain. Thirteen substitutions appeared promising and their improved antibacterial activities were confirmed for different bacteria after larger scale synthesis of these analogs. By combining two favorable substitutions into one peptide, we finally obtained an oncocin analog that was ten times more active against P. aeruginosa and even 100-fold more active against S. aureus than the original oncocin, providing minimal inhibitory concentrations of 4–8 and 0.5 µg/mL, respectively.
KeywordsOncocin Proline-rich antimicrobial peptide Pseudomonas aeruginosa Quartz crystal microbalance SPOT synthesis
We thank Laszlo Otvos for helpful discussions and proofreading. Financial support by the European Fund for Regional Structure Development (EFRE, European Union and Free State Saxony, no. 13405/2286), the Federal Ministry of Education and Research (BMBF; no. 01GU1104A), and German Academic Exchange Service (DAAD, no. 56265084) is gratefully acknowledged.
Compliance with ethical standards
This study was funded by the European Fund for Regional Structure Development (EFRE, European Union and Free State Saxony, Grant number 13405/2286), the Federal Ministry of Education and Research (BMBF, Grant number 01GU1104A), and German Academic Exchange Service (DAAD, Grant number. 56265084).
Conflict of interest
Ralf Hoffmann is a cofounder of AMP Therapeutics GmbH (Leipzig, Germany) and member of its scientific advisory board. Daniel Knappe was a part-time coworker of AMPT. The authors filed a patent application including the manuscript data under file number WO 2013/064633 (“Oncopeltus peptide derivatives as antimicrobial peptides”, May 10, 2013).
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