Structure–Activity Relationships of the Adrenocorticotropins and Melanotropins: The Synthetic Approach

Rat liver polysome N alpha-acetyltransferase has been purified to homogeneity by a four-step procedure that utilizes ammonium sulfate precipitation, gel filtration, hydroxylapatite chromatography, and Mono Q ion exchange chromatography. The enzyme is greatly stabilized by the inclusion of EDTA and 0.01% deoxycholate in the isolation buffers. The purified enzyme has a native molecular weight of 190,000 and a subunit molecular weight of 95,000, suggesting that it is a homodimer. The enzyme shows a pH optimum of 8.0 and is strongly inhibited by Cl-, I-, SCN-, and ClO4- and to a lesser degree by sulfate and acetate. It is unaffected by phosphate, citrate, and F- and by Na+ and K+; NH4+ is partially inhibitory. The enzyme is also sensitive to iodoacetic acid. It is generally more similar to yeast N alpha-acetyltransferase [Lee, F.-J. S., Lin, L.-W., & Smith, J. A. (1988) J. Biol. Chem. 263, 14948-14955] than to the hen oviduct enzyme, which contains a 7S RNA subunit [Kamitani, K., & Sakiyama, F. (1989) J. Biol. Chem. 264, 13194-13198], although the amino acid compositions are quite different.

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ACTH Antagonists

Montibeller

Finn

1974

Proc. Natl. Acad. Sci. U.S.A.

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Structural modifications within the active site of the ACTH molecule have produced analogs that inhibit the hormone sensitive adenylate cyclase system of bovine adrenal cortical plasma membranes. It is demonstrated that the tryptophan residue of the ACTH molecule is essential for stimulation of the enzyme. Substitution of tryptophan by phenylalanine or by NG-methyltryptophan as in [Gln5, provides ACTH analogs that exhibit high affinity for the ACTH receptor(s) but fail to activate the adenylate cyclase system. It is concluded that affinity for the receptors alone is not sufficient for expression of hormonal activity. The observation that adrenal cortical adenylate cyclase activated by fluoride ion is not inhibited by the antagonists indicates that hormonal and fluoride activation proceed via different mechanisms.The discovery of competitive inhibitors of polypeptide hormones is of considerable importance from a fundamental as well as from a practical point of view. Not only are compounds exhibiting such properties valuable tools for probing the mechanism of action of polypeptide hormones but they may have clinical importance in controlling specific endocrinopathies. Considerable efforts are presently directed toward discovery of such compounds. Knowledge of the essential structural features (active site) of the parent hormone may provide clues for the development of competitive antagonists.We have regarded the S-peptide-S-protein system* (1) as a model for the mode of action of peptide hormones (2). In this system two enzymically inactive components, S-peptide and S-protein, combine in a highly specific manner to form the enzymically fully active ribonuclease S. In the model, Sprotein is analogous to the receptor; S-peptide represents a peptide hormone. Structure-function studies (3) have demonstrated that [His"2] is the active site of S-peptide and that the rest of the molecule provides the vehicle to bring this residue into the correct stereochemical position on S-protein to form the active site of ribonuclease S. Since mechanistic studies (4) had shown that the pK of the imidazole ring of [His12] in Speptide is of prime importance for enzymic activity, it was a logical step to replace this histidine with an isosteric amino acid exhibiting a different pK from histidine in order to develop an S-peptide antagonist. It has now become evident that many peptide hormones combine with receptors on the cell surface and bring about stimulation of hormone sensitive adenylate cyclase(s) (6). This activation appears to provide the key to physiological function. Details of the mechanisms involved are missing but peptide hormones, like S-peptide, do indeed function as enzyme activators.The sequence -His-Phe-Arg-Trp-Gly-(positions 6-10 in Fig. 1) was postulated to be the essential structural feature (active site) of the ACTH molecule (2) and structure-function studies based on in vivo assays have largely confirmed this hypothesis (7). Based on observations with the S-peptide-S-protein system, we speculated that a...

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Structure–Activity Relationships of the Adrenocorticotropins and Melanotropins: The Synthetic Approach

Cited by 4 publications

References 187 publications

Hyperglycemic effect in the rabbit induced by ACTH4–10

Hyperglycemic effect in the rabbit induced by ACTH4–10

Rat liver polysome N.alpha.-acetyltransferase: isolation and characterization

ACTH Antagonists

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