An unusual structural motif of antimicrobial peptides containing end-to-end macrocycle and cystine-knot disulfides.
Four macrocyclic cystine-knot peptides of 29-31 residues, kalata, circulin A and B (CirA and CirB), and cyclopsychotride, have been isolated from coffee plants but have undetermined physiological functions. These macrocycles and 10 of their analogs prepared by chemical synthesis were tested against nine strains of microbes. Kalata and CirA were specific for the Gram-positive Staphylococcus aureus with a minimum inhibition concentration of approximately 0.2 microM. They were relatively ineffective against Gram-negative bacteria such as Escherichia coli and Pseudomonas aeruginosa. However, CirB and cyclopsychotride were active against both Gram-positive and Gram-negative bacteria. In particular, CirB showed potent activity against E. coli with a minimum inhibitory concentration of 0.41 microM. All four cyclic peptides were moderately active against two strains of fungi, Candida kefyr and Candida tropicalis, but were inactive against Candida albicans. These macrocycles are cytotoxic and lysed human red blood cell with a lethal dose 50% of 400 microM. Modifying the Arg residue in kalata with a keto aldehyde significantly reduced its activity against S. aureus whereas blocking the arg in CirA produced no significant effect. The two-disulfide variants and their scrambled disulfide isomers exhibited antimicrobial profiles and potency similar to their native peptides. However, in high-salt assays (100 mM NaCl), few of these macrocyclic peptides, natives or analogs, retained antimicrobial activity. These results show that the macrocyclic peptides possess specific and potent antimicrobial activity that is salt-dependent and that their initial interactions with the microbial surfaces may be electrostatic, an effect commonly found in defensin antimicrobial peptides. Furthermore, their end-to-end cyclic structure with a cystine-knot motif represents a molecular structure of antimicrobials and may provide a useful template for the design of novel peptide antibiotics.
Tam
J P
JP
Department of Microbiology and Immunology, Vanderbilt University, A-5119 MCN, 1161 21st Avenue South, Nashville, TN 37232-2363, USA. tamjp@ctrvax.vanderbilt.edu
Lu
Y A
YA
Yang
J L
JL
Chiu
K W
KW
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D000935:Antifungal Agents / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D002175:Candida / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003069:Coffee / Q000737:chemistry; D003198:Computer Simulation; D047169:Cyclotides; D003553:Cystine; D004220:Disulfides; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects; D006090:Gram-Negative Bacteria / Q000187:drug effects; D006094:Gram-Positive Bacteria / Q000187:drug effects; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010456:Peptides, Cyclic / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D011487:Protein Conformation; D011550:Pseudomonas aeruginosa / Q000187:drug effects; D013211:Staphylococcus aureus / Q000187:drug effects
D000900:Anti-Bacterial Agents; D000935:Antifungal Agents; D003069:Coffee; D047169:Cyclotides; D004220:Disulfides; D010456:Peptides, Cyclic; C093020:cyclopsychotride A; C091930:kalata B1; C102385:circulin B; D003553:Cystine; C102384:circulin A