title: Novel synthetic analogues of avian β-defensin-12: the role of charge, hydrophobicity, and disulfide bridges in biological functions.
abstract: BACKGROUND
  Avian β-defensins (AvBD) possess broad-spectrum antimicrobial, LPS neutralizing and chemotactic properties. AvBD-12 is a chemoattractant for avian immune cells and mammalian dendritic cells (JAWSII) - a unique feature that is relevant to the applications of AvBDs as chemotherapeutic agents in mammalian hosts. To identify the structural components essential to various biological functions, we have designed and evaluated seven AvBD analogues.
  
  
  RESULTS
  In the first group of analogues, the three conserved disulfide bridges were eliminated by replacing cysteines with alanine and serine residues, peptide hydrophobicity and charge were increased by changing negatively charged amino acid residues to hydrophobic (AvBD-12A1) or positively charged residues (AvBD-12A2 and AvBD-12A3). All three analogues in this group showed improved antimicrobial activity, though AvBD-12A3, with a net positive charge of +9, hydrophobicity of 40% and a predicted CCR2 binding domain, was the most potent antimicrobial peptide. AvBD-12A3 also retained more than 50% of wild type chemotactic activity. In the second group of analogues (AvBD-12A4 to AvBD-12A6), one to three disulfide bridges were removed via substitution of cysteines with isosteric amino acids. Their antimicrobial activity was compromised and chemotactic activity abolished. The third type of analogue was a hybrid that had the backbone of AvBD-12 and positively charged amino acid residues AvBD-6. The antimicrobial and chemotactic activities of the hybrid resembled that of AvBD-6 and AvBD-12, respectively.
  
  
  CONCLUSIONS
  While the net positive charge and charge distribution have a dominating effect on the antimicrobial potency of AvBDs, the three conserved disulfide bridges are essential to the chemotactic property and the maximum antimicrobial activity. Analogue AvBD-12A3 with a high net positive charge, a moderate degree of hydrophobicity and a CCR2-binding domain can serve as a template for the design of novel antimicrobial peptides with chemotactic property and salt resistance.
authors.affiliation: Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Missouri, Columbia, MO, 65211, USA.
authors.forename: Ming
authors.initials: M
authors.lastname: Yang
authors.affiliation: Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Missouri, Columbia, MO, 65211, USA.
authors.forename: Chunye
authors.initials: C
authors.lastname: Zhang
authors.affiliation: Department of Biomedical Science, College of Veterinary Medicine, University of Missouri, Columbia, MO, 65211, USA.|Veterinary Medical Diagnostic Laboratory, College of Veterinary Medicine, University of Missouri, Columbia, MO, 65211, USA.
authors.forename: Michael Z
authors.initials: MZ
authors.lastname: Zhang
authors.affiliation: Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Missouri, Columbia, MO, 65211, USA. zhangshup@missouri.edu.|Veterinary Medical Diagnostic Laboratory, College of Veterinary Medicine, University of Missouri, Columbia, MO, 65211, USA. zhangshup@missouri.edu.
authors.forename: Shuping
authors.initials: S
authors.lastname: Zhang
mesh_terms: D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D000890:Anti-Infective Agents / Q000138:chemical synthesis / Q000494:pharmacology; D001419:Bacteria / Q000187:drug effects / Q000254:growth & development; D058491:Bacterial Load; D001717:Birds / Q000276:immunology*; D002460:Cell Line / Q000187:drug effects; D002633:Chemotaxis; D002645:Chickens; D015169:Colony Count, Microbial / Q000379:methods; D003545:Cysteine / Q000737:chemistry; D003713:Dendritic Cells / Q000276:immunology; D004220:Disulfides / Q000737:chemistry*; D004338:Drug Combinations; D057927:Hydrophobic and Hydrophilic Interactions; D008264:Macrophages / Q000187:drug effects; D008826:Microbial Sensitivity Tests / Q000379:methods; D008855:Microscopy, Electron, Scanning; D010455:Peptides / Q000138:chemical synthesis; D011487:Protein Conformation; D012694:Serine / Q000737:chemistry; D012965:Sodium Chloride / Q000008:administration & dosage / Q000494:pharmacology; D023083:beta-Defensins / Q000008:administration & dosage / Q000138:chemical synthesis* / Q000494:pharmacology*
chemical_list: D000890:Anti-Infective Agents; D004220:Disulfides; D004338:Drug Combinations; D010455:Peptides; D023083:beta-Defensins; D012965:Sodium Chloride; D012694:Serine; D003545:Cysteine; D000409:Alanine
keywords: Antimicrobial activity; Avian β-defensin analogues; Chemotactic activity; Disulfide bridges; Hydrophobicity; Net positive charge