Abstrakt

Aims: The aim of this work was to characterize oligopeptide uptake in Candida albicans mutants in which the genes encoding putative peptide permeases were selectively disrupted. Initial velocities of transport of model oligopeptides, (Ala)2, (Ala)3, (Ala)4, and sensitivity of mutants to antifungal oligopeptides containing N3-(4-methoxyfumaroyl)-L-2,3-diaminopropanoic acid (FMDP) were determined and compared to those of the wild-type SC5314 strain.Methods: C. albicans mutants lacking one or a set of the genes encoding putative oligopeptide permeases (opt1Δ, opt 23Δ, opt4Δ, opt5Δ, opt6Δ, opt7Δ, opt8Δ, opt12345Δ), the genes coding for di-/tripeptide permeases (ptr222Δ), or all peptide permeases (opt12345ptr222Δ) were constructed in the laboratory of one of the co-authors (JM). Oligopeptides containing FMDP (Nva-FMDP, Lys-Nva-FMDP and (Met)3-FMDP) were synthesized at Gdańsk University of Technology. Peptide uptake was followed using the colorimetric method of determination of peptide concentration employing 2,4,6-trinitrobenzenesulfonate. MIC determination was done using the twofold serial dilution method according to the EUCAST requirements. Additionally, MICs were determined in YNB medium containing different nitrogen sources.Results: All tested mutant strains, except for ptr222Δ and opt12345ptr222Δ, were similarly sensitive to Nva-FMDP and Lys-Nva-FMDP. A correlation between growth inhibitory activity and initial transport velocity was observed. The opt12345ptr222Δ mutant was resistant to FMDP-peptides and unable to transport all tested model peptides. The ptr222Δ mutant was unable to transport (Ala)2 and (Ala)3 and sensitive to (Met)3-FMDP, whereas the opt1Δ mutant was unable to transport (Ala)4 and resistant to the antifungal tetrapeptide. The MIC values of FMDP-oligopeptides highly depended on medium composition. Strong differences between the mutants were noticed in the case of sensitivity to (Met)3-FMDP in YNB medium containing ammonium sulfate as a nitrogen source. An interesting observation, yet to be fully explained, was that the ptr222Δ mutant was sensitive to Lys-Nva-FMDP in YNB medium supplemented with sodium glutamate.Conclusions: The OPT1 gene encodes the oligopeptide permease of major importance. Genes encoding the PTR permeases, PTR2 and PTR22, are essential for the uptake of di- and tripeptides and the sensitivity to antifungal peptides of the same size.

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