No abstract

Human cystatin C (HCC) is one of the amyloidogenic proteins to be shown to oligomerize via a three-dimensional domain swapping mechanism. This process precedes the formation of a stable dimer and proceeds particularly easily in the case of the L68Q mutant. According to the proposed mechanism, dimerization of the HCC precedes conformational changes within the beta2 and beta3 strands. In this article, we present conformational studies, using circular dichroism and MD methods, of the beta2-L1-beta3 (His43-Thr72) fragment of the HCC involved in HCC dimer formation. We also carried out studies of the beta2-L1-beta3 peptide, in which the Val57 residue was replaced by residues promoting beta-turn structure formation (Asp, Asn, or Pro). The present study established that point mutation could modify the structure of the L1 loop in the beta-hairpin peptide. Our results showed that the L1 loop in the peptide excised from human cystatin C is broader than that in cystatin C. In the HCC protein, broadening of the L1 loop together with the unfavorable L68Q mutation in the hydrophobic pocket could be a force sufficient to cause the partial unfolding and then the opening of HCC or its L68Q mutant structure for further dimerization. We presume further that the Asp57 and Asn57 mutations in the L1 loop of HCC could stabilize the closed form of HCC, whereas the Pro57 mutation could lead to the opening of the HCC structure and then to dimer/oligomer formation.

Modifications in angiotensin II and its antagonistic peptides that should have increased in vivo half-lives but not reduced biological activity were studied by determining the effect of a-methylation of the tyrosine in position 4. angiotensin II, synthesized by the solid-phase procedure, showed 92.6 ± 5.3% pressor activity of angiotensin H. Incubation with a-chymotrypsin for 1 hr indicated absence of degradation although, under the same conditions, angiotensin H was completely degraded to two components. Comparison of the 'H NMR spectra in aqueous solution and the circular dichroism spectra in trifluoroethanol of angiotensin II and [a-methyltyrosine-4]angiotensin II suggested that a methylation of the tyrosine residue in angiotensin HI does not lead to major changes in the overall solution conformation. These results are in contrast to those obtained with N-methylation in position 4, which drastically reduced the biological activity and produced remarkable changes in the peptide backbone and a severe limitation in rotational freedom of the side chains in tyrosine. Thus, it may be possible to synthesize potent angiotensin II analogs that have greater resistance to enzymatic degradation by a-methylation in position 4 (or 5) and simultaneous suitable modification at the NH2 and COOH termini.The antagonists for the pressor hormone angiotensin II (AspArg-Val-Tyr-Ile-His-Pro-Phe; All) have proved useful in the study ofexperimental hypertension and as possible new clinical diagnostic tools (1). However, their long-term application has been limited because they have a short in vivo half-life and, due to degradation by peptidases, are not orally active. We therefore attempted to make these peptides resistant to this enzymatic degradation by replacing the natural amino acid residues with N-methylamino (2) or /3homoamino (3) acid residues. These modifications drastically reduced the biological activity ofthese analogs, and conformation studies suggest that N-methylation in positions 4 or 5 results in remarkable changes in the peptide backbone and a severe limitation in the rotational freedom of the side chains in tyrosine, isoleucine, and histidine residues (4). The replacement ofan a proton by an a methyl group, however, is thought to produce minimal changes in backbone and side-chain structures. The analogs thus obtained might mimic the parent hormone in recognizing and binding with the receptor on the cell membrane and, in addition, be stable to enzymatic degradation (4, 5). Based on this hypothesis, we have synthesized [a-methyltyrosine4]angiotensin II ([a-MeTyr4]AII).And, indeed, this peptide is resistant to chymotrypsin degradation and yet retains almost the full pressor activity of All. Conformation studies suggest minimum changes in backbone and side-chain structures. These results again suggest that, for full biological activity (agonist or antagonist), the backbone and side-chain structure of the analog should resemble that of the hormone, All (2, 4). These observations may be helpful in the design of new...

The following derivatives of 7-hydroxytetrahydroisoquinoline-3-carboxylic acid {Tic(OH) [I]}, a conformationally restricted analogue of tyrosine, were synthesized for the purpose of photophysical studies and in order to elucidate the nature of tyrosine fluorescence and its decay:, Ac-Ala-Tic(OH) [VI], and Tic(OH)-Gly-NH 2 [VII]. For the simple Tic(OH) derivatives I-IV, the N-methylamide was found to be a more effective quencher than the acetyl group. For the peptidic derivatives V-VII the highest quenching of the fluorescence of the phenolic chromophore was observed in the case of Ala-Tic(OH). The simple Tic(OH) derivatives I-IV were also the subject of theoretical studies (MOPAC 93). The obtained thermodynamic parameters (MOPAC calculations) and the fluorescence components were discussed on the basis of the rotamer theory in order to explain the participation of an individual rotamer in the complex process of the fluorescence decay of tyrosine.

No abstract