Holger Wenschuh

Protected amino acid chlorides vs protected amino acid fluorides: Reactivity comparisons.

Abstract Although Fmoc amino acid fluorides are excellent reagents for coupling of moderately hindered amino acids ( e.g. , Aib-to-Aib) they are not suited for significantly more hindered systems ( e.g. , Aib-to-MeAib). While urethane-protected acid chlorides are inherently more reactive than the fluorides they are also ineffective for hindered systems due to competing oxazolone formation. This limitation is by-passed if urethane protection is replaced by arenesulfonyl protection and the Aib-to-MeAib and even the MeAib-to-MeAib couplings are easily achieved via the appropriate acid chlorides but not the acid fluorides.

Proline at position 14 of alamethicin is essential for hemolytic activity, catecholamine secretion from chromaffin cells and enhanced metabolic activity in endothelial cells

Alamethicin is known to lyse different biological cells and to induce voltage dependent ion channels in lipid bilayers. A set of analogs with proline shifted from position 14 in the native peptide towards the N- and C-terminus was used to investigate the role of proline in: (i) alamethicin induced hemolysis of human red blood cells, (ii) stimulation of catecholamine secretion from bovine adrenal chromaffin cells and (iii) induction of metabolic activity in bovine aortic endothelial cells. Half maximal hemolytic activity was found at 30 microM alamethicin concentration, complete lysis occurred at 100 microM. The stimulation of catecholamine secretion in the presence of extracellular Ca2+ was concentration dependent up to 50 microM alamethicin. At this high concentration mild secretion was also found in the absence of Ca2+ indicating cell membrane damage. Alamethicin transiently stimulated the metabolic rate of endothelial cells in a concentration dependent mode up to 20 microM while the inhibition of metabolism at higher concentrations pointed to a toxic effect. The alamethicin analogs were completely inactive in all the biological assays. The effects correlated with a loss of dye release inducing activities on phosphatidylcholine vesicles and reduction of channel forming properties in lipid bilayers and were associated with modifications of membrane affinity rather than conformational changes of the peptides. The results indicate that proline at position 14 of the native peptide is essential for the interaction with different membrane systems.

The ease of peptide detection by matrix-assisted laser desorption ionization mass spectrometry: the effect of secondary structure on signal intensity

Several structurally well-characterized model peptides were used to examine the relationship between peptide structure and signal intensity in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). It was found that peptides displaying stable alpha-helical and beta-sheet structures show lower signal intensities than the corresponding analogs having disturbed secondary structures caused by substitution of two adjacent amino acids by their D isomers. Since such substitutions do not affect properties other than the secondary structure propensity, the differences observed are ascribed to this phenomenon or some related effect such as association. The results indicate that the formation of stable secondary structures in peptides may be a possible source of incomplete peptide mass fingerprints resulting from protein digestion and for difficulties in the quantitative evaluation of peptide mixtures via MALDI-MS.