Makoto Tamaki

Fatty acyl-gramicidin S derivatives with both high antibiotic activity and low hemolytic activity

In the present study, novel eight GS derivatives having the octanoyl-(Lys)(n)- moieties, cyclo{-Val-Orn-Leu-d-Phe-Pro(4β-NH-X)-Val-Orn-Leu-d-Phe-Pro-} {X=-H (1), and -(Lys)(n)-CO(CH(2))(6)CH(3)n=0 (2), 1 (3), 2 (4), and 3 (5)} and cyclo{-Val-Orn-Leu-d-Phe-Pro(4α-NH-X)-Val-Orn-Leu-d-Phe-Pro-} {X=-H (6), and -(Lys)(n)-CO(CH(2))(6)CH(3)n=1 (7), and 2 (8)} were synthesized. Among them, 4, 5 and 8 result the high antibiotic activity against both Gram-positive and Gram-negative microorganisms tested. In addition, 4 and 5 showed very low hemolytic activity compared with that of GS. Thus, the introduction of the excess amino groups and the fatty acyl moiety to the γ-NH(2) group of Pro(5) residue in GS molecule lowered the unwanted hemolytic activity and enhanced the desired antibiotic activity.

A Novel, Antimicrobially Active Analog of Gramicidin S without Amphiphilic Conformation

A novel gramicidin S analog, cyclo(-Val-Leu-Leu-Orn-Leu-D-Phe-Pro-)2, was synthesized, its antibiotic activity compared with gramicidin S and shown to be as potent as gramicidin S when compared with the susceptibility toward five Gram-positive microorganisms. It exceeded the activity of gramicidin S against Bacillus megaterium ATCC 19213 by a factor of two. Circular dichroism and NMR data suggested this analog to adopt an antiparallel β-sheet conformation without amphiphilic character.

Design and syntheses of gramicidin S analogs, cyclo(-X-Leu-X-D-Phe-Pro-) 2 (X=His, Lys, Orn, Dab and Dap)

Design and syntheses of gramicidin S analogs, cyclo(-X-Leu-X-D-Phe-Pro-) 2 (X=His, Lys, Orn, Dab and Dap)

A Novel Active Analogue of Gramicidin S with Smaller Ring Size

A novel active gramicidin S analogue with smaller ring size, cyclo[-δ-Orn(-Val-Pro-d-Phe-H)-Leu-]2, was synthesized and examined the structure-activity relationship. Its analogue showed antibiotic activity against all Gram-positive microorganisms tested, and its activity was 1/2∼1/8 of that of gramicidin S. The present results indicated that both structures of cyclo(-δ-Orn-Leu-)2 and H-d-Phe-Pro-Val sequence play the important role for showing the antibiotic activity.

Studies of gramicidin S analogues having various ring sizes by reversed-phase high-performance liquid chromatography

Many gramicidin S analogues containing six to fourteen amino acid residues were synthesized and their chromatographic behaviour was investigated using reversed-phase high-performance liquid chromatography. Cyclo(-Val-Leu-Orn-Leu-D-Phe-Pro-Val-Orn-Leu-D-Phe-Pro-) gave double peaks in the chromatogram, whereas the other peptides gave a single peak. The influences of the concentration of this peptide, column temperature and flow-rate on the chromatographic separation were examined. The isomeric conformers were separated from each of the double peaks and were in equilibrium with each other at low temperatures. These results suggested that the presence of the additional L-Leu residue preceding the Orn residue gives rise to moderate stabilization of their conformers.

Solvent-dependent conformations of a cyclic tetrapeptide

The conformations of cyclo(-γ-Abu-L-Pro-)2 in several solvents have been characterized by means of 1H and 13C NMR, CD and IR spectra. CD results indicate that the change in polarity of the solvent induces an inversion of the conformation, revealed by the disappearance of the positive peak near 210 nm and then by the gradual appearance of a negative trough near 210 nm having opposite chirality. The data of NMR spectroscopy in CDCl3, CD2Cl2, CD3OH, (CD3)2SO and D2O indicate that cyclo(-γ-Abu-L-Pro-)2 has a C2 symmetric conformation consisting of the cis–trans–cis–trans peptide bond backbone (with two cis γ-Abu-L-Pro bonds) in all solvents, and that the change of polarity of solvent induces the inversion of cis and trans conformations around the Pro αC–CO single bond. (The trans and cis regions describe the rotational states of the Pro αC–CO single bond in which the Pro αC–H is trans and cis to the direction of the Pro carbonyl oxygen atom, respectively). The cis–trans–cis–trans conformation containing the two cis Pro αC–CO bonds in CDCl3 and CD2Cl2 is stabilized by the presence of seven-membered rings consisting of intramolecular hydrogen bonds between γNH and αCO in the γ-Abu residue.

A novel polycationic analogue of gratisin with antibiotic activity against both Gram-positive and Gram-negative bacteria.

AbstractA novel polycationic analogue of gratisin, cyclo(-Val-Orn-Leu-D-Phe-Pro-D-Lys-)2, was designed and synthesized, which exhibited strong activity against all Gram-positive and Gram-negative bacteria tested. Its activity against Pseudomonas aeruginosa IFO 3080 was two times higher than gramicidin S.

Biomimetic synthesis of a peptide antibiotic, gratisin

Recently, we reported the direct formation of a peptide antibiotic, gramicidin S (GS), cyclo(-D-Phe-Pro-Val-Orn-Leu-)2, by the dimerization–cyclization of pentapeptide active esters, D-Phe-Pro-Val-Orn-Leu-ONSu (-ONSu: succinimide ester), having the sequence identical with that of the linear precursor pentapeptide in the biosynthesis of GS and no protecting group on the side-chain of the Orn residue. This biomimetic approach has been extended to the synthesis of a peptide antibiotic, gratisin (GR), cyclo(-D-Phe-Pro-D-Tyr-Val-Orn-Leu-)2, isolated from Bacillus brevis Y-33. In the cyclization of six hexapeptide succinimide esters having a Val, Orn, Leu, D-Phe, Pro or D-Tyr residue at each C-terminus, only H-D-Phe-Pro-D-Tyr-Val-Orn-Leu-ONSu gave semi-GR and GR in yields of 31 and 8%, respectively. Other hexapeptide esters did not give semi-GR and GR. In both biomimetic syntheses of GS and GR, the amino acid sequences having a Leu residue at the C-terminus are essential. In addition, a change in the concentration of peptide and the polarity of reaction solvents influenced greatly the yields of cyclic monomer (semi-GR) and cyclic dimer (GR). However, the yield of GR by dimerization–cyclization of hexapeptide active ester was lower when compared with the direct formation of GS (38%). The difference may result from differences in the chain length and the configurations of amino acid residues around the Pro residue.

Studies on cyclic peptides related to gratisin by reversed-phase high-performance liquid chromatography.

The chromatographic behaviour of several synthetic peptides related to the antibiotic peptide gratisin was investigated using reversed-phase high-performance liquid chromatography. The influence of the concentration of the peptides, column temperature and flow-rate on the chromatographic separation was examined. Of these peptides, some analogues with the D-X-D-Y-L-Pro or L-Pro-D-X-D-Y sequences and strong activities gave double peaks in the chromatogram. The isomeric conformers were separated from each of the double peaks and were in equilibrium with each other at low temperatures. It is proposed that the presence of the sequences of D-X-D-Y-L-Pro and L-Pro-D-X-D-Y in these peptides gives rise to the stabilization and various degrees of hydrophobicity of their conformers.