Amino acid sequences, renin

Affinity labeling ATP sites, 194-96 creatine kinase, 200-205 Amastatin, 94-96 Amide bond hydrolysis, 227 Amino acid sequences, renin inhibitors, 139,141f D-Amlno acids and activity,... 

In this work we will focus on the use of the cubic phase as a delivery system for oligopeptides - Desmopressin, Lysine Vasopressin, Somatostatin and the Renin inhibitor H214/03. The amino acid sequences of these peptides are given in Table I. The work focuses on the cubic phase as a subcutaneous or intramuscular depot for extended release of peptide drugs, and as a vehicle for peptide uptake in the Gl-tract. Several examples of how the peptide drugs interact with this lipid-water system will be given in terms of phase behaviour, peptide self-diffusion, in vitro and in vivo release kinetics, and the ability of the cubic phase to protect peptides from enzymatic degradation in vitro. Part of this work has been described elsewhere (4-6). 

A number of chemical approaches have been used in the design of renin inhibitors. In the absence of the purified enzyme, most of the early search for inhibitors was carried out using crude renin preparations. The amino acid sequences... 

Figure 1. Amino acid sequences of renin inhibitors based on the amino acid sequence of angiotensinogen. Details of the synthesis and assay are given in references 11, 13, and 14.

Chemistry of the renin-angiotensin system. The amino acid sequence of the amino terminal of human angiotensinogen is shown. R denotes the remainder of the protein molecule. See text for additional steps in the formation and metabolism of angiotensin peptides. 

A number of chemical approaches have been used in the design of renin inhibitors. In the absence of the purified enzyme, most of the early search for inhibitors was carried out using crude renin preparations. The amino acid sequences of mouse, rat and human renin were obtained later on using either the traditional isolation and sequencing techniques or cDNA methodology. Various three-dimensional models of renin were constructed in the early stages, based on the x ray structures of other similar aspartyl proteases, for example endothia-pepsin and penicillopepsin. Later on, the X ray crystal structure of recombinant human renin was reported. The inhibitor design process has been based on some of these models. 

Renin-angiotensin system. R is the remainder of the amino acid sequence. 

Analysis of the amino acid sequences of PLE isoenzymes revealed that the remarkably small differences of ca. 20 amino acids are not distributed randomly but are located within distinct conserved areas. Among the different isoenzymes, PLE-1 and an isoenzyme termed A-PLE ° [247] were shown to be most useful for stereoselective ester hydrolysis. The latter enzyme, which was expressed at a high level in Pichia pastoris, is remarkably stable and showed perfect enantioselectivity for the resolution of methyl (4 )-5-chloro-2-isopropyl-4-pentenoate, which is a key building block for the synthesis of the renin inhibitor aliskiren, which is used in the treatment of hypertension (Scheme 2.36) [248]. 

Comparison of SQ 20,881 with fifty-seven related synthetic peptides indicated that the last five amino acids of the sequence are required for significant enzyme inhibiting activity (100). This nonapeptide, intravenously, lowered blood pressure even in patients with normal renin levels (101. 102). This effect is strongly augmented by sodium depletion. 

The strategy of designing saquinavir was based on the transition-state mimetic concept, an approach that has been used successfully in the design of potent inhibitors of renin and other aspartic proteases [10]. From the variety of nonscissile transition-state analogs of a dipeptide, the hydroxyethylamine mimetic was selected because it most readily accommodates the amino acid moiety characteristic of the Phe-Pro and Tyr-Pro cleavage sequence of the... 

The Pro-Phe amide linkage was used as a seed fragment for GROW, which was run in unrestricted mode (i.e., it was permitted to choose, at each position in the sequence, any one of the 20 naturally occurring amino acids). Of the sequences produced, that with the highest score was Ac-Pro-Trp-Trp-Phe-Arg-Arg-NH2 (the Ac-Pro fragment being retained from the inhibitor mentioned above). When synthesized and assayed, this compound was found to inhibit renin with a K-, of 30 xM. 

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