Design of nontoxic analogues of cathelicidin-derived bovine antimicrobial peptide BMAP-27: the role of leucine as well as phenylalanine zipper sequences in determining its toxicity.
BMAP-27 is a cathelicidin-derived bovine antimicrobial peptide, which shows moderate cytotoxicity and potent antibacterial activity against a wide variety of microorganisms. Despite a number of studies, very little is known about the amino acid sequences of this peptide that controls its antibacterial and cytotoxic activities. Small stretches of phenylalanine and leucine zipper sequences were identified at the N- and C-termini of the molecule, respectively. To understand the structural and functional roles of these sequence elements, we synthesized and characterized several analogues of BMAP-27 after substituting leucine or phenylalanine residue(s) at a and/or d positions of the leucine and phenylalanine zipper sequences, respectively, with alanine. BMAP-27 analogues exhibited significantly reduced cytotoxicity against the human red blood (hRBC) and murine 3T3 cells as compared to that of the wild-type peptide. Interestingly, BMAP-27 and its analogues exhibited comparable antibacterial activity against the selected Gram-positive and Gram-negative bacteria. Moreover, BMAP-27 and its analogues exhibited similar localization and assembly onto the selected bacteria and induced comparable permeability in these cells. However, only BMAP-27, not its analogues, assembled and bound strongly onto the hRBCs and permeabilized them. The results indicated that not only a leucine zipper but also a phenylalanine zipper sequence plays an important role in maintaining the assembly of BMAP-27 on the mammalian cells examined here and cytotoxic activity against them. To the best of our knowledge, this is the first report of the evaluation of structural and functional roles of a phenylalanine zipper sequence in a naturally occurring antimicrobial peptide.
Molecular and Structural Biology Division, Central Drug Research Institute, CSIR,Lucknow 226001, India.
Aqeel
A
Ahmad
Sarfuddin
S
Azmi
Raghvendra M
RM
Srivastava
Saurabh
S
Srivastava
Brijesh K
BK
Pandey
Rubha
R
Saxena
Virendra Kumar
VK
Bajpai
Jimut Kanti
JK
Ghosh
D016475:3T3 Cells; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000502:physiology; D000817:Anilino Naphthalenesulfonates / Q000737:chemistry; D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry* / Q000494:pharmacology / Q000633:toxicity*; D001419:Bacteria / Q000166:cytology / Q000187:drug effects; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D015195:Drug Design; D031541:Fluorescence Resonance Energy Transfer; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D016350:Leucine Zippers / Q000502:physiology*; D008081:Liposomes / Q000737:chemistry; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology / Q000633:toxicity; D010539:Permeability / Q000187:drug effects; D010649:Phenylalanine / Q000378:metabolism*; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry* / Q000494:pharmacology / Q000633:toxicity*; D013050:Spectrometry, Fluorescence
D000817:Anilino Naphthalenesulfonates; D023181:Antimicrobial Cationic Peptides; C102359:BMAP-27; D008081:Liposomes; D010455:Peptides; D011506:Proteins; D010649:Phenylalanine; C027132:1-anilino-8-naphthalenesulfonate