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    Cyclotide-rich fractions containing nanofibers by electrospinning: preparation, characterization and examination of antimicrobial activity
    (Tubitak Scientific & Technological Research Council Turkey, 2022) Bas, Elif Busra; Kurekc, Cemil; Kalfa, Orhan Murat; Akcan, Muharrem
    In this study, antimicrobial nanofibers were produced with the mixtures of polyvinyl alcohol (PVA) and cyclotide-rich fractions by electrospinning method. After extraction, the first separation was carried out with C18 flash chromatography and then fractioned into five separate parts by reversed-phase high-pressure liquid chromatography (RP-HPLC). The molecular weights of cyclotides in each fraction were determined by quadrupole time-of-flight liquid chromatography-mass spectrometry (Q-TOF LC-MS). Cyclotide-rich fractions were mixed with 10% of PVA solution and nanofibers were produced from this biocomposite mixture by electrospinning method. The nanofibers were characterized by field emission scanning electron microscopy (FE-SEM), and it was observed that 100% peptide-containing nanofibers (cyclotide-rich fraction/10% PVA, w/v) had more regular fiber textures. The presence of the peptides in the nanofiber was also confirmed by analytical RP-HPLC, as the peptides in both peptide fractions and nanofiber solutions have the same retention times. The nanofibers produced with the fourth cyclotide-rich fraction showed activity against gram-positive bacteria (Bacillus cereus) in antimicrobial susceptibility test. As a result of these findings, cyclotide-containing nanofibers with antimicrobial activity can be produced for pharmaceutical research and development studies.
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    Design and modification of frog skin peptide brevinin-1GHa with enhanced antimicrobial activity on Gram-positive bacterial strains
    (Springer Wien, 2022) Kara, Seyda; Kurekci, Cemil; Akcan, Muharrem
    Naturally occurring frog skin peptides are one of largest sources of antimicrobial peptides that have many advantages including high potency, broad spectrum of targets and low susceptibility to multiple drug-resistance bacteria. However, they also have disadvantages such as hemolytic activity, low stability and high production costs. For these reasons, various strategies have been applied to overcome these drawbacks restricting their use in clinical trials. Previously reported brevinin-1GHa (BR-1GHa) is a 24 amino acid long antimicrobial peptide isolated from Hylarana guentheri with hemolytic activity. To enhance the antimicrobial activity of this peptide and to reduce its hemolytic activity, we designed five new temporin like analogues and examined their bioactivities. Temporins are another class of frog skin peptides without hemolytic activity and shorter than brevinins. When the antimicrobial activities of new analogues were examined against a panel of microorganisms, BR-1GHa-3, in which two alanine residues in the truncated version of BR-1GHa were replaced with leucine, exhibited significantly improved antimicrobial activity against Gram-positive bacterial strains (e.g., S. aureus ATCC 29213 and E. casseliflavus ATCC 700327) with lower hemolytic activity compared to the BR-1GHa peptide. Furthermore, BR-1GHa-4 analogue, in which Gly3 was replaced with Pro, did not show any hemolytic activity except for highest (128 mu M) concentration tested and have a strong antimicrobial effect on Gram-positive bacteria (e.g., E. faecalis ATCC 51299 and B. cereus ATCC 13061).