title: Single Amino Acid Substitutions at Specific Positions of the Heptad Repeat Sequence of Piscidin-1 Yielded Novel Analogs That Show Low Cytotoxicity and In Vitro and In Vivo Antiendotoxin Activity.
abstract: Piscidin-1 possesses significant antimicrobial and cytotoxic activities. To recognize the primary amino acid sequence(s) in piscidin-1 that could be important for its biological activity, a long heptad repeat sequence located in the region from amino acids 2 to 19 was identified. To comprehend the possible role of this motif, six analogs of piscidin-1 were designed by selectively replacing a single isoleucine residue at a d (5th) position or at an a (9th or 16th) position with either an alanine or a valine residue. Two more analogs, namely, I5F,F6A-piscidin-1 and V12I-piscidin-1, were designed for investigating the effect of interchanging an alanine residue at a d position with an adjacent phenylalanine residue and replacing a valine residue with an isoleucine residue at another d position of the heptad repeat of piscidin-1, respectively. Single alanine-substituted analogs exhibited significantly reduced cytotoxicity against mammalian cells compared with that of piscidin-1 but appreciably retained the antibacterial and antiendotoxin activities of piscidin-1. All the single valine-substituted piscidin-1 analogs and I5F,F6A-piscidin-1 showed cytotoxicity greater than that of the corresponding alanine-substituted analogs, antibacterial activity marginally greater than or similar to that of the corresponding alanine-substituted analogs, and also antiendotoxin activity superior to that of the corresponding alanine-substituted analogs. Interestingly, among these peptides, V12I-piscidin-1 showed the highest cytotoxicity and antibacterial and antiendotoxin activities. Lipopolysaccharide (12 mg/kg of body weight)-treated mice, further treated with I16A-piscidin-1, the piscidin-1 analog with the highest therapeutic index, at a single dose of 1 or 2 mg/kg of body weight, showed 80 and 100% survival, respectively. Structural and functional characterization of these peptides revealed the basis of their biological activity and demonstrated that nontoxic piscidin-1 analogs with significant antimicrobial and antiendotoxin activities can be designed by incorporating single alanine substitutions in the piscidin-1 heptad repeat.
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: Amit
authors.initials: A
authors.lastname: Kumar
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: Amit Kumar
authors.initials: AK
authors.lastname: Tripathi
authors.affiliation: Electron Microscopy Unit, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: Manoj
authors.initials: M
authors.lastname: Kathuria
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: Sonal
authors.initials: S
authors.lastname: Shree
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: Jitendra Kumar
authors.initials: JK
authors.lastname: Tripathi
authors.affiliation: Sophisticated Analytical Instrumentation Facility, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: R K
authors.initials: RK
authors.lastname: Purshottam
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow, India Academy of Scientific and Innovative Research, Chennai, India.
authors.forename: Ravishankar
authors.initials: R
authors.lastname: Ramachandran
authors.affiliation: Electron Microscopy Unit, CSIR-Central Drug Research Institute, Lucknow, India Academy of Scientific and Innovative Research, Chennai, India.
authors.forename: Kalyan
authors.initials: K
authors.lastname: Mitra
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow, India Academy of Scientific and Innovative Research, Chennai, India jk_ghosh@cdri.res.in.
authors.forename: Jimut Kanti
authors.initials: JK
authors.lastname: Ghosh
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000890:Anti-Infective Agents / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D023181:Antimicrobial Cationic Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000187:drug effects; D015195:Drug Design; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects; D005260:Female; D029941:Fish Proteins / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D006090:Gram-Negative Bacteria / Q000187:drug effects / Q000254:growth & development; D006094:Gram-Positive Bacteria / Q000187:drug effects / Q000254:growth & development; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology / Q000187:drug effects; D051379:Mice; D008807:Mice, Inbred BALB C; D008826:Microbial Sensitivity Tests; D010538:Peritonitis / Q000139:chemically induced / Q000401:mortality / Q000473:pathology / Q000517:prevention & control*; D015202:Protein Engineering; D017433:Protein Structure, Secondary; D013329:Structure-Activity Relationship; D016019:Survival Analysis
chemical_list: D000890:Anti-Infective Agents; D000894:Anti-Inflammatory Agents, Non-Steroidal; D023181:Antimicrobial Cationic Peptides; D029941:Fish Proteins; D008070:Lipopolysaccharides; C449802:moronecidin protein, Morone saxatilis