Alanine gramicidin

Other workers began to study the structure of gramicidin. Christensen and coworkers12 isolated crystalline tryptophane and leucine from a hydrolysate. They found no evidence for a fatty acid component and established that phenylalanine, proline and hydroxyproline were absent from a hydrolysate. These workers isolated alanine diox-pyridate from a hydrolysate and also established that gramicidin contained a compound with vicinal hydroxy and amino groups. They speculated that this compound might be serine or isoserine and proposed that gramicidin contains two tryptophane, 2 leucine, 2 or 3 alanine and 1 hydroxyamino residues or a multiple of this composition. 

Gordon, Martin and Synge18 utilized their new and elegant technique, chromatography, to establish the amino acid composition of gramicidin. They proposed a 24 unit cyclic peptide consisting of six moles each of leucine, and tryptophane, 5 moles of valine, 3 moles of alanine and 2 moles each of glycine and unknown hydroxyamino compound. 

A non-ribosomal biosynthetic pathway is clearly indicated for cyclosporin A, considering the uncommon structural elements MeBmt, L-a-aminobutyric acid and D-alanine as well as the plethora of isolated congeners [20,21]. Non-ribosomal biosynthesis directed by multienzyme thiotemplates have been reported for other small peptides of microbial origin, for example, gramicidin S [22] and enniatin [23]. Experimental data for cyclosporin A were obtained by feeding appropriate labelled precursors to cultures of T. inflation strains. The distribution profile of the labelled atoms in cyclosporin A was determined by 3H- or 13C-NMR spectroscopy. In preliminary trials with several tritium and carbon-14 labelled precursors, [met/y>/-3H]methionine proved to be the most suitable marker for the biosynthetic preparation of radiolabelled cyclosporin A for pharmacokinetic and metabolic studies [24],... 

Gramicidin A, however, is attacked by JV -bromoacetamide (NBA) and N-bromosuccinimide (NBS) (Gross and Witkop, unpublished observation). In 50 % aqueous ethyl alcohol at room temperature 5 % of the peptide bonds (20% of the tryptophyl peptide bonds) are cleaved with NBS. Methyl alcohol must be avoided because it opens the spirodioxindole lactone from oxidized tryptophan to the ester even at room temperature. The cleavage mixture separates on electrophoresis (pH = 2.5, sodium-formate buffer) into four ninhydrin-positive components of which the fastest migrating one was identified as ethanolamine. Dinitrophenylation showed leucine and alanine to be additional NHs-terminals of the released fragments. 

A nonribosomal biosynthetic pathway is clearly indicated for CyA, considering its cyclic structure, the partial N methylation of the 11 peptide bonds, the uncommon building units Bmt, L l-aminobutyric acid, and D-alanine, and the plethora of isolated congeners. Nonribosomal biosynthesis directed by multieniyme thiotemplates have been reported for other small peptides of microbial origin, such as gramicidin S and enniacin (for a review, see Ref. 39). 

Ala = alanine DMPC = dimyristoyl phosphatidylcholine GA = gramicidin A Gly = glycine Leu = leucine Phe = phenylalanine PRC = photosynthetic reaction center SDS = sodium dodecyl sulfate Trp = tryptophan Tyr = tyrosine Val = valine. 

Antibiotics belong to very diverse chemical classes of compoimds. Azaserine and chloramphenicol have already been mentioned (Chapt. II-2). Another peptidic antibiotic is gramicidin S which contains an amino acid of the rare mseries, D-phenyl-alanine. It is a cyclic peptide with ten amino acids ... 

The lability of tryptophan under the conditions usually employed for acid hydrolysis of proteins (6N HCl, 24 hrs, in sealed tubes under vacuum) is not due to instability of the indole nucleus under such conditions, but to side reactions involving non-proteinaceus material, like carbohydrates, or to the presence of particular amino acids in the acidic hydrolysis mixture. In fact, gramicidin A, a pentadecapeptide containing, apart from tryptophan, only purely aliphatic amino acids like glycine, alanine, leucine and valine, gives a quantitative recovery of tryptophan after acid hydrolysis in 6N HCl (l20). 

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