Synthesis of thiazole, imidazole and oxazole containing amino acids for peptide backbone modification.

Novel 5-ring heterocyclic building blocks are synthesized. These can be incorporated into analogs of peptide antibiotics such as microcin B17, which is a potent DNA-gyrase inhibitor that exhibits eight thiazole and oxazole moieties. In particular, the syntheses of imidazole and bisoxazole amino acids as novel peptidomimetics are reported, this includes a new procedure for the oxidative conversion of the intermediates oxazoline, imidazoline as well as oxazole-oxazoline into the corresponding heteroaromatic compounds. A mixture of 1,8-diazabicyclo-[5.4.0.]-undec-7-ene carbon tetrachloride/acetonitrile and pyridine proved to be a very effective and mild agent.

Synthesis and antimicrobial activity in vitro of new amino acids and peptides containing thiazole and oxazole moieties.

2-(Pyrrolidinyl)thiazole-4-carboxylic acid 5d, 2-(1-aminoalkyl)thiazole-4-carboxamides and hydrazides 8, 10 have been synthesized using alanine, valine, and proline as educts. In addition oxazole amino acids derived from leucine 20a and alanine 20b and some peptides 13, 14, 16 containing the 5-ring heterocyclic backbone modifications have been prepared. The thiazole and oxazole containing amino acids and peptides showed moderate antibacterial activity in vitro against various Gram-positive (Staphylococcus aureus, Bacillus cereus, etc.) and Gram-negative (Escherichia coli, Protens vulgaris, etc.) bacteria, fungi (Candida albicans), and yeast (Saccharomyces cerevisae, etc.).

Structure-affinity studies of C-terminally modified analogs of neuropeptide Y led to a novel class of peptidic Y1 receptor antagonist.

A novel type of C-terminally modified analogs of the 36-mer peptide hormone neuropeptide Y has been synthesized, characterized and tested with respect to receptor affinity and biological activity in various systems. The compounds were obtained by synthesizing the fully protected peptide fragment NPY 1-35 or analogs of this, and coupling it in solution to various amines, alcohols, and modified tyrosine residues. It could be confirmed, that the C-terminal tyrosineamide of NPY is essential for its affinity to the Y1 receptor subtype. Obviously, the amino group of the amide part is more important than the oxygene atom of the carbonyl group, as NPY 1-35-tyrosinol has a lower affinity than NPY 1-35-tyrosinethioamide. NPY 1-35-tyramide could be shown to act as an antagonist in a Ca2+ release assay in human neuroblastoma cells. Analogs of NPY 1-35-tyramide showed the same structure-affinity relationships as NPY itself, suggesting, that there exists the same binding mode for the agonist and the antagonist.

Identification of receptor binding sites by competitive peptide mapping: phages T1, T5, and phi 80 and colicin M bind to the gating loop of FhuA.

Previously we proposed a transmembrane model of the FhuA receptor protein in the outer membrane of Escherichia coli. Removal of the largest loop at the cell surface converted the FhuA transport protein into an open channel and rendered cells resistant to the FhuA-specific phages T1, T5, and phi 80 and to colicin M. In the present study we employed acetylated hexapeptide amides covering the entire surface loop to investigate binding of the phages and of colicin M. Competitive peptide mapping proved to be a powerful technique to uncover three ligand binding sites within a region of 34 amino acid residues. Hexapeptides derived from three specific regions of the surface loop inhibited infection of cells by the phages and killing by colicin M. Two of these regions were common among all four FhuA ligands. Electron microscopy of phage T5 revealed that one inhibitory peptide triggered a strong conformational change leading to the release of DNA from the phage head. These results suggest that the FhuA gating loop is the target for specific binding of phages T1, T5, and phi 80 and colicin M.

Studies on the total synthesis of an A7,B7-dicarbainsulin. III. Assembly of segments and generation of biological activity.

As a further contribution to the synthesis of an insulin analogue with a stable A7-B7 interchain bond, the synthesis of A(8-21) by solution methods, and of B(9-25) as well as [7-(2,7-diaminosuberic acid)]B(1-8) by solid phase methods is described. In the latter compound, the amino group of the diaminosuberic acid residue was acylated with A(1-6), and the resulting "U-peptide" sequentially elongated with the C-terminal A- and finally B-chain sequences. The conversion of the product into the disulfide moiety gave a mixture which could not be resolved by currently available methods. However, the low biological activity of the crude product indicates that the A7-B7 disulfide bond is not crucially important for the activity of insulin.

Synthesis of A7,B7-dicarbainsulin, an analogue with a noncleavable bond between A- and B-chain. II. Synthesis of the A-chain segments.

As part of the total synthesis of [A7,B7-L,L-2,7-diaminosuberoyl]-des-(B26-B30)-insulin B25-amide, an insulin analogue containing a non-cleavable bond between A- and B-chain, the chemical synthesis of the A-chain segments is described. The N-terminal sequence A(1-6), Boc-Gly-Ile-Val-Glu(OBut)-Gln-Cys(SBut)-NH-NH2, was synthesized in solution. The middle segment A(8-16), Ddz-Thr(But)-Ser(But)-Ile-Cys(SBut)-Ser(But)-Leu-Tyr- (But)-Gln-Leu-NH-NH2, was obtained by solid phase synthesis according to the Fmoc strategy. The C-terminal segment A(17-21), Bpoc-Glu(OBut)-Asn-Tyr-Cys(Acm)-Asn-OBut, was prepared in solution.