Endomorphin-1 (Tyr-Pro-Trp-Phe-NH P ) is a highly selective and potent agonist of the W W-opioid receptor. To identify structural attributes unique to this opioid peptide and potential sites of recognition, a conformational analysis has been performed using multidimensional NMR and molecular modeling techniques. The spectroscopic results, derived from experiments in both DMSO and water, indicate that endomorphin-1 exists in the cis-and trans-configuration with respect to the Pro-omega bond in approximately 25% and 75% populations, respectively. In DMSO, the cis-configuration adopts a compact sandwich conformation in which the Tyr and Trp aromatic rings pack against the proline ring, whereas the trans-configuration adopts an extended conformation. Although non-random structure was not observed in water, condensed phase molecular dynamics calculations indicate that trans-isomers dominate the population in this higher dielectric medium. Structural comparison of the cis-and trans-configurations with morphine and selective W Wpeptide ligands PL-017 and D-TIPP, as well as the N N-selective peptide ligands TIPP (N N-antagonist, W W-agonist) and DPDPE were also performed and suggest the trans-isomer is likely the bioactive form. A hypothesis is proposed to explain W W-and N Nselectivity based on the presence of spatially distinct selectivity pockets among these ligands.z 1998 Federation of European Biochemical Societies.

Collagen, the most abundant protein in mammals, has fascinated scientists because of its extraordinary structural features and biological importance. In 1961, Rich and Crick suggested that collagen possesses a triple-helical structure, 1 which was later confirmed by X-ray analysis of 30-mer collagen-related peptides (CRPs), such as (Pro-Hyp-Gly) 4 Pro-Hyp-Ala-(Pro-Hyp-Gly) 5 and (Pro-Pro-Gly) 10 . 2 The large triple-helical domains of collagen consist of three peptide strands with Gly-X-Y repeating motifs, 300 nm in length (vs ∼9 nm for CRPs), with the X and Y mainly populated by Pro and Hyp, respectively. In addition, there are short telopeptide regions at the N-and C-termini, which are important for fibril assembly. 3 The rigidity of the ropelike super-helix and the assembled fibril helps provide mechanical strength to tissues, such as skin, tendons, ligaments, and blood vessels. Following vascular injury, the exposed collagen in the vessel wall promotes tissue repair by activating platelets for aggregation and adhesion. However, excessive platelet activity, such as from rupture of an atherosclerotic plaque, can lead to pathological thrombosis with attendant arterial obstruction, as in myocardial infarction or stroke. 4 The study of the structure, stability, and function of collagen triple helices has been facilitated by the use of synthetic collagen model peptides. 5 Although oligomerized CRPs, via dendrimer assembly or covalent crosslinking, can effectively induce platelet aggregation, less organized CRPs have lacked this property. 6 We sought to identify a single-stranded CRP that could spontaneously self-assemble into a bioactive material. Recently, two research groups obtained micrometer-scale CRP-based materials from the self-assembly of covalently attached triple-stranded entities by employing a cysteine knot. 7 We report on the design, synthesis, and characterization of novel single-stranded CRP 1a (Figure 1a), which self-assembles by noncovalent interactions into a triplehelical, micrometer-scale, composite fibrillar substance with noteworthy biological activity.The requirement of the telopeptide regions of collagen, 3 and specific Tyr and Phe residues within the C-terminal telopeptide chain, 8 suggests the importance of aromatic residues in the "code" for collagen self-assembly. 8 Inspired by nature, we sought to explore short, single-stranded CRPs that possess the main repeat-unit of collagen, Gly-Pro-Hyp, and contain aromatic recognition units at the N-and C-termini. Phenyl and pentafluorophenyl end-groups were considered, given the strong noncovalent aromatic-stacking interaction between benzene and hexafluorobenzene. 9 In this regard, 32-mer peptide 1a, with pentafluorophenylalanine (F 5 -Phe) and Phe residues at the N-and C-termini, respectively, was of interest. We speculated that interstrand aromatic-stacking and ordered hydro-

The novel findings are that obese subjects have lower levels of IPA, a solely bacterially derived tryptophan derivative, and IPA improved intestinal barrier function in vitro and DIO mice. Reduced plasma IPA levels and reversal by surgery may be a consequence of intestinal indole-producing microbiota but underlying mechanisms warrant further investigation.