Delineation of an Active Fragment and Poly(l-proline) II Conformation for Candidacidal Activity of Bactenecin 5

Abstract

Bactenecin 5 and its fragments [BN22 (1-22), BN16 (7-22), and BC24 (20-43)] were synthesized by solid-phase methods. Their antifungal activities on Candida albicans have been studied and compared with those of the native bactenecin 5. The conformational preferences of these peptides in aqueous and nonaqueous solutions and in lipid vesicles were examined by circular dichroism. The highly active N-terminal fragment (BN16) was examined in aqueous solution using 500 MHz two-dimensional NMR. Bactenecin 5 and its fragments are potent candidacidal agents against C. albicans. The N-terminal fragments (BN22 and BN16) of bactenecin 5 are relatively more active than the C-terminal fragment BC24, especially at lower concentrations. The N-terminal region (7-22) which retains the activity of the whole molecule appears to be the functional domain for candidacidal activity. The CD spectra of bactenecin 5 and its fragments are reminiscent of the CD spectrum of poly(L-proline) type II structure in aqueous and nonaqueous solutions and also in lipid vesicles. The temperature dependence of NH chemical shifts and 1H/2H exchange effect on amide resonances suggest the absence of intramolecularly hydrogen-bonded NH groups. The coupling constant (JNH-CalphaH) values, conformational restriction offered by the Pro residues (phi = -60 degrees +/- 15 degrees), the set of medium- and short-range nuclear Overhauser effects observed for the active N-terminal fragment (BN16), and the restrained structure calculation using DIANA suggest that poly(L-proline) type II conformers of the peptide molecules could be significantly populated in aqueous solution. The ability of bactenecin peptides to induce disruption of lipid vesicles correlates well with their activity. Our results suggest that poly(L-proline) type II structure may, indeed, be the biologically active conformation for candidacidal activity of bactenecin peptides.

Distinct Antifungal Mechanisms: β-Defensins RequireCandida albicansSsa1 Protein, while Trk1p Mediates Activity of Cysteine-Free Cationic Peptides

“…Peptide purity was assessed by amino acid analysis and mass spectroscopy. The bactenecin 16 (BN 16) peptide from the N terminus of bovine bactenecin was synthesized as previously described (22). HNP-1 was purchased from Sigma.…”

Susceptibility of oral bacteria to an antimicrobial decapeptide

“…Identification of most of the spin systems was achieved unambiguously from amide proton-relayed TOCSY connectivities. Assignment of resonances to individual amino acids was accomplished by combined analyses of the NH-C AE H connectivities in the fingerprint regions in the TOCSY and 2D NOE spectra, as described previously (Raj et al, 1996(Raj et al, , 1998(Raj et al, , 2000a.…”

Diversity in penaeidin antimicrobial peptide form and function

“…In contrast to apidaecin-like peptides from insects, Bac5 exhibited some level of antifungal activity apparently affiliated with the N-terminal segment of this anti-MP [18]. Moreover, the structure induced by the position of proline residues appears to strongly influence the range of activity of Bac5 [18]. The membrane penetrating properties of bactenecins are not only interesting because of the obvious effect on targeted microbes, but also because they confer the ability to penetrate other types of biological membranes.…”

“…Bac5 and Bac7) formed of repetitive motifs of proline, cationic (Arg and His) and hydrophobic (Leu, Ile, Phe and Tyr) amino acids [16,17]. In contrast to apidaecin-like peptides from insects, Bac5 exhibited some level of antifungal activity apparently affiliated with the N-terminal segment of this anti-MP [18]. Moreover, the structure induced by the position of proline residues appears to strongly influence the range of activity of Bac5 [18].…”