Rao S. Rapaka

Vacuum ultraviolet circular dichroism spectrum of β-turn in solution

The vacuum ultraviolet circular dichroism spectrum of an isolated 4 → 1 hydrogen bonded β-turn is reported. The observed spectrum of N-acetyl-Pro-Gly-Leu-OH at − 40°C in trifluoroethanol is in good agreement with the theoretically calculated CD spectrum of the β-turn conformation. This spectrum, particularly the presence of a strong negative band around 180 nm and a large ratio [θ]201/[θ]225, can be taken as a characteristic feature of the isolated β-turn conformation. These CD spectral features can thus be used to distinguish the β-turn conformation from the β-structure in solution.

Inhibition of hydroxyproline synthesis by palladium ions

Palladium ions, administered as PdSO4, markedly affect the incorporation of L-[3,4-3H2] proline into non-dialyzable fractions in 10-day chick embryo cartilage explants with a 55-65% reduction in the concentration range 0.06-0.6 mM. Under these conditions the synthesis of [3H]hydroxyproline was nearly completely inhibited. Experiments with prolyl hydroxylase (EC 1.14.11.2) indicated a strong irreversible inhibition of the enzyme with a competition between Fe2+ and Pd2+. The Ki for the inhibition was 0.02 mM. Pd2+-treated enzyme remained inactive after extensive dialysis. These studies suggest that Pd2+ may inhibit collagen synthesis by replacing Fe2+ in the active site of prolyl hydroxylase and forming strong complexes with the enzyme. These studies also point to a potential mechanism of Pd2+ toxicity.

Strategies in the racemization-free synthesis of polytripeptide models of collagen.

Synthesis of racemization-free sequential polypeptides is a major challenge to the peptide chemist. The loss of optical integrity at a single residue can drastically alter the properties of the polymer. We have investigated racemization in the synthesis of polytripeptide models of collagen. The conformational features of collagen are determined by a large imino content and the presence of Gly in every third position. Thus polymers of tripeptides containing Pro and Gly with an asymmetric residue A permit evaluation of the contribution of A to collagen conformation. in the synthesis of such polymers, the position of Pro in the tripeptide is of importance because of its variable reactivity at the N or C terminal residue. N-terminal Pro is more reactive but also sterically hinders polycondensation. Terminal Pro also participates in undesirable side reactions such as formation of urea or diketopiperazine derivatives and cyclization. Cyclization can be avoided by using concentrated solutions of monomer. Racemization occurs during the synthesis of OCl5Ph or ONp esters but not in the synthesis of ONSu esters. This, however, does not guarantee formation of optically pure products. Best results are obtained with ONSu esters and limited use of base during polymerization. Polymerization appears to be stereoselective since the largest molecular weight polymers are purest, smallest molecular weight products highly racemized.

Conformational Properties of Polypeptide Models of Collagen

Individual collagen chains exist as Polypro II helices because of their large imino content and their super-coiling into the collagen triple helix is facilitated by Gly in every third position. Because of this, collagen may be considered as being made up of Gly-led triplets. One fourth of such triplets in collagen have the sequence Gly-Pro-X and another one fifth, Gly-X-Hyp, where X is an α-amino residue. The stereochemical properties of the imino peptide bond, the position of the imino residue in the sequence and its interactions with neighboring residues, determine the conformation. Synthetic polytripeptides of sequence (Gly-Pro-X)n generate collaqen-like conformations in aqueous solutions whereas (Gly-X-Pro)n sequences usually do not. This difference has been attributed to different H-bonding properties of X-NH in the two sequences. Stabilizing interactions are said to occur between the side chain of X and Pro ring atoms in X-Pro but not in Pro-X. We investigated this question using (Gly-Pro-Sar)n (I) and (Gly-Sar-Pro)n (II). (I) generated collagen-like conformations in aqueous solutions but (II) did not although some order was elicited in helix promoting solvents. Since NH-H bonds are not possible at Sar residues, they may not play a final role in stabilizing the collagen helix. It appears likely that non-bonding interactions of the imino residue with the residue on its C-terminal may play a significant role in stabilizing collagen-like conformations.