DNA transcription is initiated by a small regulatory region of transactivators known as the transactivation domain. In contrast to the rapid progress made on the functional aspect of this promiscuous domain, its structural feature is still poorly characterized. Here, our multidimensional NMR study reveals that an unbound fulllength p53 transactivation domain, although similar to the recently discovered group of loosely folded proteins in that it does not have tertiary structure, is nevertheless populated by an amphipathic helix and two nascent turns.

A high resolution structure of ␣-conotoxin EI has been determined by 1 H NMR spectroscopy and molecular modeling. ␣-Conotoxin EI has the same disulfide framework as ␣4/7 conotoxins targeting neuronal nicotinic acetylcholine receptors but antagonizes the neuromuscular receptor as do the ␣3/5 and ␣A conotoxins. The unique binding preference of ␣-conotoxin EI to the ␣ 1 /␦ subunit interface of Torpedo neuromuscular receptor makes it a valuable structural template for superposition of various ␣-conotoxins possessing distinct receptor subtype specificities. Structural comparison of ␣-conotoxin EI with the ␥-subunit favoring ␣-conotoxin GI suggests that the Torpedo ␦-subunit preference of the former originates from its second loop. Superposition of three-dimensional structures of seven ␣-conotoxins reveals that the estimated size of the toxin-binding pocket in nicotinic acetylcholine receptor is ϳ20 Å (height) ؋ 20 Å (width) ؋ 15 Å (thickness).The nicotinic acetylcholine receptors (nAChRs) 1 are a well studied family of ligand-gated ion channels comprising a diverse set of molecular subtypes (1). The best characterized is the receptor at the neuromuscular junction, with four different subunits in a pentameric array, i.e. (␣ 1 ) 2 ␤ 1␥ ␦. Less well understood and more diverse are the neuronal nAChRs that assemble in exogenous expression systems with a general composition of (␣ m ) 2 (␤ n ) 3 , where m ϭ 2-6 and n ϭ 2-4, or (␣ 7 ) 5 (2, 3). A large variety of ligands bind to such diverse nAChRs via unknown mechanisms. A better understanding of the ligandbinding mechanism would be possible if a suitable three-dimensional structure of nAChR were available. Although cryoelectron microscopic images of nAChR show the spatial arrangement of five subunits of the receptor and some aspects of the ligand-binding pockets (4, 5), they are as yet insufficient for describing ligand-receptor interactions in atomic detail. In this regard, the recently determined crystal structure of acetylcholine-binding protein is expected to provide useful insight into ligand-nAChR interactions (6).Small peptide toxins of Conus origin known as the ␣-and ␣A-conotoxins are highly useful tools for exploring ligandnAChR interactions (7,8). A well defined subgroup of ␣-conotoxins, referred to as the ␣3/5 2 subfamily conotoxins (Table I), are antagonists of neuromuscular receptors and show, except for ␣-conotoxin SI (9, 10), binding preference to the ␣ 1 /␥ subunit interface of Torpedo nAChR. On the other hand, more recently found ␣A-conotoxins differ from the ␣3/5 subfamily conotoxins in the amino acid sequences (11, 12) as well as in their threedimensional structures (13). The ␣A-conotoxins, nevertheless, target neuromuscular receptors. A third subfamily of ␣-conotoxins known as the ␣4/7 subfamily has also been found; members of this subfamily target subtypes of neuronal and muscle nAChRs (14 -17). The ␣4/7 subfamily contains two disulfide bonds like the ␣3/5 subfamily but has a different spacing between the disulfide bonds.Even though a high resolutio...

We report the solution three-dimensional structure of an ␣A-conotoxin EIVA determined by nuclear magnetic resonance spectroscopy and restrained molecular dynamics. The ␣A-conotoxin EIVA consists of 30 amino acids representing the largest peptide among the ␣/␣A-family conotoxins discovered so far and targets the neuromuscular nicotinic acetylcholine receptor with high affinity. ␣A-Conotoxin EIVA consists of three distinct structural domains. The first domain is mainly composed of the Cys 3 -Cys 11 -disulfide loop and is structurally ill-defined with a large backbone root mean square deviation of 1.91 Å. The second domain formed by residues His 12 -Hyp 21 is extremely well defined with a backbone root mean square deviation of 0.52 Å, thus forming a sturdy stem for the entire molecule. The third C-terminal domain formed by residues Hyp 22 -Gly 29 shows an intermediate structural order having a backbone root mean square deviation of 1.04 Å. A structurally ill-defined N-terminal first loop domain connected to a rigid central molecular stem seems to be the general structural feature of the ␣A-conotoxin subfamily. A detailed structural comparison between ␣A-conotoxin EIVA and ␣A-conotoxin PIVA suggests that the higher receptor affinity of ␣A-conotoxin EIVA than ␣A-conotoxin PIVA might originate from different steric disposition and charge distribution in the second loop "handle" motif.The fish-hunting cone snail Conus ermineus is the only known piscivorous Conus species in the Atlantic Ocean (1, 2). In aquaria, the species captures its prey by extending its highly distensible amber-colored proboscis from which ejected is a hollow harpoon-shaped tooth that functions as a hypodermic needle to inject venom into the fish. Venom injection results in a complete inhibition of neuromuscular transmission in the injected prey. A major molecular component of this physiological strategy is to prevent neurotransmitter (acetylcholine) binding to the major postsynaptic receptor, the muscle subtype of the nicotinic acetylcholine receptor.In most fish-hunting cone snail venoms, the competitive nicotinic antagonists of the skeletal muscle subtype are peptides with two disulfide bonds belonging to the ␣-conotoxin family (3). However, in C. ermineus, there are two different nicotinic receptor antagonists: 1) ␣-conotoxin EI with two disulfide bonds (4) and 2) ␣A-conotoxin EIVA with three disulfide bonds (5). The ␣A-conotoxins have been found in only two Conus species: 1) C. ermineus from the Atlantic and 2) the eastern Pacific purple cone, Conus purprascens (6). These species are believed to have a different evolutionary history from the Indo-Pacific fish-hunting Conus (7,8).Highly selective antagonistic activity against particular subtypes of nicotinic acetylcholine receptors (nAChRs) 1 (Table I) has rendered ␣/␣A-conotoxins an important tool for studying molecular function of nAChR (3, 9 -11). In an effort to discover novel modulators of nAChR by identifying critical residues within the toxin for receptor contact, we have been applying a...

In the absence of an experimental bTLR8 structure, recent studies have called attention to the fact that bTLR8 can also be activated by hTLR7/hTLR8 agonist, such as antiviral imidazoquinoline derivatives of resiquimod (R848) and imiquimod (R837) as well as some guanine nucleotide analogs with a scaffold structure related to the nucleic acids of ssRNA virus. In particular, the known small agonists (namely CL075, CL097 and R848) have been targeted to determine distinguishable deciding factors in complex with dimeric bTLR8-ECDs in comparison to ligand-induced activated hTLR8-ECDs. According to basic knowledge, the deciding eligibility criteria can be subsequently applied in our bTLR8 model to characterize the 3D-arrangement of chemical features (pharmacophore) and to investigate the distinct restrictions affecting species-specificity on dual TLR7/TLR8 small agonists suggested in previous works. Despite the lack of extensive structural biology studies regarding the interaction of bTLR8-ECDs with the agonists, our complex models of bTLR8-ECDs and the known agonists were applied to identify the deciding factors required for the interactions from agonist-based and (bTLR8-agonist complexes) structure-based pharmacophores. These pharmacophore constraints impose their essential chemical features to active bTLR8 receptors. The characterized pharmacophores all were employed in the virtual screening of candidates with a further acting factor of calf immune enhancer. Two hits were suggested as satisfying all decision factors to identify a potent bTLR8-specific agent with novel scaffolds dissimilar to imidazoquinoline analogues lacking overall homogeneity.

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