ABSTRACTa-Melanocyte-stimulating hormone (a-MSH) reversibly darkens frog skins by stimulating melanosome movement (dispersion) within melanophores. Heat-alkali treatment of a-MSH results in prolonged biological activity of the hormone. Quantitative gas chromatographic analysis of the hydrolyzed heat-alkali-treated peptide revealed partial racemization particularly at the 4 (methionine) and 7(phenylalanine) a-Melanotropin (a-MSH, a-melanocyte-stimulating hormone) is a tridecapeptide (Ac-Ser-Tyr-Ser-Met-Glu-His-Phe-ArgTrp-Gly-Lys-Pro-Val-NH2) that is synthesized and secreted by the pars intermedia of the vertebrate pituitary (1). The amino acid residues that are important in the expression of melanotropic activity have been elucidated through systematic structure-function investigations of a-MSH and a-MSH fragments on amphibian melanophores (2, 3) and, to a lesser extent, on mammalian melanoma cells (4-6). Very little information is available, however, regarding the stereochemical and conformational correlates of biological activity in either of these two biological systems.Earlier reports have shown that heat-alkali treatment of crude or purified preparations of naturally occurring a-MSH produces a partially racemized product with altered activity on amphibian melanophores both in vivo and in vitro. Such changes in biological effects have been discussed in terms of "potentiation," "prolongation," and "retardation" (7-12). Although the precise biochemical mechanism by which these unusual biological properties were produced is unknown, it appeared possible that synthetic stereostructural tailoring of a-MSH might produce an analogue that would also possess these properties. Utilizing a high-resolution gas chromatographic method to localize and quantitate specific sites of racemization within the primary sequences of peptides, we obtained additional evidence which suggested that stereochemical substitution at position 7 (replacement of L-phenylalanine by D-phenylalanine) of a-MSH or [Nle4]-a-MSH would provide an analogue with the desired biological properties. Previous investigations have shown that [Nle4]-a-MSH is more potent than a-MSH on both amphibian melanophores (2, 6) and on stimulating melanoma adenylate cyclase (6, 13), and it is also resistant to inactivation by chloramine-T (14, 15), an oxidant used in peptide iodination. Because heat-alkali treatment of this analogue also resulted in "potentiation," "prolongation," and "retardation," it was clear that alteration of the methionine residue was not a requirement for the expression of these properties. Thus, it was decided to retain the benefits of the norleucine substitution in position 4 in the synthesis of the "definitive" peptide.We report here the synthesis of [Nle4, D-Phe7]-a-MSH and present data demonstrating its unique biological properties. These include prolonged biological activity, enhanced potency relative to a-MSH in a number of biological systems, and resistance to degradation by serum enzymes. The biological properties of this analogue provide...
Many amino acids contain an asymmetric centre, occurring as laevorotatory, L, or dextrorotatory, D, compounds. It is generally assumed that abiotic synthesis of amino acids on the early Earth resulted in racemic mixtures (L- and D-enantiomers in equal abundance). But the origin of life required, owing to conformational constraints, the almost exclusive selection of either L- or D-enantiomers, and the question of why living systems on the Earth consist of L-enantiomers rather than D-enantiomers is unresolved. A substantial fraction of the organic compounds on the early Earth may have been derived from comet and meteorite impacts. It has been reported previously that amino acids in the Murchison meteorite exhibit an excess of L-enantiomers, raising the possibility that a similar excess was present in the initial inventory of organic compounds on the Earth. The stable carbon isotope compositions of individual amino acids in Murchison support an extraterrestrial origin -- rather than a terrestrial overprint of biological amino acids-although reservations have persisted. Here we show that individual amino-acid enantiomers from Murchison are enriched in 15N relative to their terrestrial counterparts, so confirming an extraterrestrial source for an L-enantiomer excess in the Solar System that may predate the origin of life on the Earth.
The membrane-bound glycoprotein dipeptidyl peptidase IV (DP IV, CD26) is a unique multifunctional protein, acting as receptor, binding and proteolytic molecule. We have determined the sequence and 1.8 Å crystal structure of native DP IV prepared from porcine kidney. The crystal structure reveals a 2-2-2 symmetric tetrameric assembly which depends on the natively glycosylated -propeller blade IV. The crystal structure indicates that tetramerization of DP IV is a key mechanism to regulate its interaction with other components. Each subunit comprises two structural domains, the N-terminal eight-bladed -propeller with open Velcro topology and the C-terminal ␣͞-hydrolase domain. Analogy with the structurally related POP and tricorn protease suggests that substrates access the buried active site through the -propeller tunnel while products leave the active site through a separate side exit. A dipeptide mimicking inhibitor complexed to the active site discloses key determinants for substrate recognition, including a Glu-Glu motif that distinguishes DP IV as an aminopeptidase and an oxyanion trap that binds and activates the P 2-carbonyl oxygen necessary for efficient postproline cleavage. We discuss active and nonactive site-directed inhibition strategies of this pharmaceutical target protein.serine protease ͉ oxyanion hole ͉ substrate channeling ͉ drug design ͉ diabetes mellitus
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