Recombinant DNA technology has now made it possible to produce proteins for pharmaceutical applications Consequently, proteins produced via biotechnology now comprise a significant portion of the drugs currently under development Isolation, purification, formulation, and delivery of proteins represent significant challenges to pharmaceutical scientists, as proteins possess unique chemical and physical properties These properties pose difficult stability problems A summary of both chemical and physical decomposition pathways for proteins is given Chemical instability can include proteolysis, deamidation, oxidation, racemization, and β-elimination Physical instability refers to processes such as aggregation, precipitation, denaturation, and adsorption to surfaces Current methodology to stabilize proteins is presented, including additives, excipients, chemical modification, and the use of site-directed mutagenesis to produce a more stable protein species

Stability of Protein Pharmaceuticals

Organic Synthesis on Soluble Polymer Supports: Liquid-Phase Methodologies.

It is a privilege to be able to contribute to this volume in which Professor Zervas’ friends, students, and colleagues have joined together to honor him and his remarkable contributions to peptide chemistry. Although I did not know him until after his days in the Bergmann Laboratory at the Rockefeller Institute, I can claim the distinction of now working in those very same rooms that they occupied back in the mid-1930s. Like all peptide chemists, I am greatly indebted to Professor Zervas, having depended so heavily on the carbobenzoxy group and on the various modified urethan protecting groups which have been direct extensions of the revolutionary advance that Bergmann and Zervas made.

Solid-phase peptide synthesis.

Soluble polymers as scaffolds for recoverable catalysts and reagents.

Candida lipolytica synthetisiert das Antibioticum Tryptanthrin aus 1 Mol Tryptophan und 1 Mol Anthranilsaure. Bei Verfutterung von Tryptophan und substituierter Anthranilsaure, bzw. substituiertem Tryptophan und Anthranilsaure, konnten die zu erwartenden Tryptanthrinderivate isoliert und identifiziert werden. Die Enzyme der Tryptanthrinbiosynthese wiesen in bezug auf diese Substrate, mit Ausnahme von Bromtryptophan, keine Spezifitat auf. Parallel zu diesen Versuchen wurden durch chemische Synthese substituierte Tryptanthrine hergestellt. Die Dierivate wurden auf ihre antibiotische Wirksamkeit gepruft; als besonders wirksam erwiesen sich die halogenierten Verbindungen.

Stoffwechselprodukte von Mikroorganismen

Kinetic studies of the liquid phase peptide synthesis.

Die Liquid‐Phase‐Methode zur Peptidsynthese; Strategie und experimentelle Grundlagen

The conformation of polyethylene glycol-bound peptides, synthesized by the liquid-phase method, was investigated. This marcromolecular C-terminal protecting group is transparent in the visible and the ultraviolet range to 190 nm and solubilizes peptides in many different solvents. The CD spectra of the polymer-bound myoglobin sequence 66–73 and of the biologically active undecapeptide “substance P” were measured in each step of the synthesis. In both examples the formation of a secondary structure during the growth of the peptide chain was found.



In the hydrophobic octapeptide containing the myoglobin sequence 66–73, the influence of either the blocked or the free N-terminal amino group on the conformation was observed. The blocked octapeptide in trifluoroethanol showed a higher degree of α-helix contribution than in its free state.



The conformation of the polyethylene glycol-bound nona- and decaalanine in trifluoroethanol and water was determined. The peptide with a free amino end group has β-conformation in trifluoroethanol as well as in water. The corresponding N-Boc-protected derivatives show helical structure. The amino end group has a decisive influence on the formation of β-structure.



The method of CD investigation of polymer-bound peptide sequences during the peptide synthesis in solution enables one to determine the influence of protecting groups and the chain end of a peptide on its conformation. It is also possible to study the relationship between the secondary structure, the chain length, and the kinetic of the coupling reaction in different solvents. Since the crystallization method for the liquid-phase peptide synthesis allows one to synthesize peptides in very short time, a new method of studying peptide conformations is opened.

[Conformation studies on oligoalanines, substance P, and the myoglobin sequence (66-73). Circular dichroism of polyethyleneglycol-bound peptides].

Fluorine-19 nuclear magnetic resonance for the control of peptide synthesis

Aus Laboratoriumskulturen eines Dermatophyten wurde eine Substanz isoliert, die Pilze (Paecilomyces varioti, Mucor miehei), nicht aber Bakterien (Bacillus subtilis, Escherichia coli) hemmt. Die antibiotische Wirkung wird durch L-Histidin und L-Threonin aufgehoben. Die Struktur des Antibioticums wurde durch gaschromatographische und massenspektrometrische Untersuchungen als L-Argnyl-D-allo-threonyl-L-phenylalanin erkannt und durch Vergleich mit dem synthetischen Tripeptid L-Arginyl-DL-allo-threonyl-L-phenylalanin bestatigt. Synthetisches L-Arg-L-Thr-L-Phe ist antifungisch inaktiv. Bedingungen fur Fermentation und Gewinnung des Antibioticums werden angegeben.



Metabolic Products of Microorganisms, 117. L-Arginyl-D-allo-threonyl-L-phenylalanine, an Amino Acid Antagonist from the Fungus Keratinophyton terreum



An antibiotically active tripeptide was isolated from laboratory cultures of the dermatophyte Keratinophyton terreum. Inhibitory effects have been observed on fungi (Paecilomyces varioti, Mucor miehei) but not on bacteria (Bacillus subtilis, Escherichia coll). The inhibition is antagonized by L-histidine and by L-threonine. The structure was elucidated by gas chromatographic and mass spectrometric investigations and was confirmed by comparison with synthetic L-arg-DL-allo-thr-L-phe. Synthetic L-arg-L-thr-L-phe does not show antifungal activity. Conditions for the fermentation and isolation of the antibiotic are reported.

Rainer Uhmann

Papers

Kinetic studies of the liquid phase peptide synthesis.

Die Liquid‐Phase‐Methode zur Peptidsynthese; Strategie und experimentelle Grundlagen

[Conformation studies on oligoalanines, substance P, and the myoglobin sequence (66-73). Circular dichroism of polyethyleneglycol-bound peptides].

Fluorine-19 nuclear magnetic resonance for the control of peptide synthesis

Stoffwechselprodukte von Mikroorganismen, 117. L‐Arginyl‐D‐allo‐threonyl‐L‐phenylalanin, ein Aminosäure‐Antagonist aus dem Pilz Keratinophytonterreum