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.

The neuronal nicotinic acetylcholine receptors constitute a highly diverse group, with subtypes consisting of pentameric combinations of ␣ and ␤ subunits. ␣-Conotoxins are a homologous series of small peptides that antagonize these receptors. We present the three-dimensional solution structure of ␣-conotoxin AuIB, the first 15-residue ␣-conotoxin known to selectively block the ␣ 3 ␤ 4 nicotinic acetylcholine receptor subtype. The pairwise backbone and heavy-atom root mean square deviation for an ensemble of 20 structures are 0.269 and 0.720 Å, respectively. The overall fold of ␣-conotoxin AuIB closely resembles that of the ␣4/7 subfamily ␣-conotoxins. However, the absence of Tyr 15 , normally present in other ␣4/7 members, results in tight bending of the backbone at the C terminus and effectively renders Asp 14 to assume the spatial location of Tyr 15 present in other neuronal ␣4/7 ␣-conotoxins. Structural comparison of ␣-conotoxin AuIB with the ␣ 3 ␤ 2 subtype-specific ␣-conotoxin MII shows different electrostatic surface charge distributions, which may be important in differential receptor subtype recognition.The ␣-conotoxins are small neuropharmacologically active peptides of Conus origin that antagonize the nicotinic acetylcholine receptor (nAChR) 1 (1). The natural diversity of biosynthesized conotoxin peptides has led to the classification of a wide spectrum of disulfide-bridged peptides, which attack various ligand and ion-gated channels and receptors (1). The nicotinic acetylcholine receptors exhibit considerable diversity in their own right because of the different compositions found in the pentameric subunits constituting each nAChR subtype (2). Although the mammalian neuromuscular subtype comprises (␣ 1 ) 2 ␤␥␦ or (␣ 1 ) 2 ␤⑀␦ subunits, the neuronal subtypes are comparatively more diverse with their hetero-or homopentameric combinations of ␣ (␣ 2 ϳ ␣ 9 ) or ␤ (␤ 2 ϳ ␤ 4 ) subunits (3, 4). The general conotoxin strategy of diversification is "combinatorial" (5), in which amino acid residues are varied within a given disulfide framework to specifically and selectively bind various subtypes of the target channel or receptor. For the case of ␣-conotoxins, target selectivity is essentially defined depending on which subunit interface of the nAChR (e.g. ␣ 1 /␥, ␣ 1 /␦, and ␣ 3 /␤ 2 ) each individual ␣-conotoxin preferentially binds to (1). Highly selective ␣-conotoxins that permit differential blocking of diverse nAChR subtypes have served as effective tools in studying these receptors (1).Of recent particular interest are the ␣-conotoxins that act on neuronal nAChRs. For example, ␣-conotoxin AuIB specifically targets the ␣ 3 ␤ 4 subtype (6), whereas ␣-conotoxin MII selectively blocks the ␣ 3 ␤ 2 subtype (7). On the other hand, ␣-conotoxin ImI, the smallest of all ␣-conotoxins and distinct because of its ␣4/3 disulfide framework 2 , is a specific antagonist of the homomeric ␣ 7 subtype (8). In addition, other neuronal ␣-conotoxins such as PnIA (9), PnIB (9), and EpI (10) that are less selective to a particu...

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...

A combination of methodologies using the extremely high mass accuracy and resolution of 15-T Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry (MS) was introduced for the identification of intact cancer cell phospholipids. Lipids from a malignant glioma cell line were initially analyzed at a resolution of >200,000 and identified by setting the mass tolerance to ±1 mDa using matrix-assisted laser desorption/ionization (MALDI) 15-T FT-ICR MS in positive ion mode. In most cases, a database search of potential lipid candidates using the exact masses of the lipids yielded only one possible chemical composition. Extremely high mass accuracy (<0.1 ppm) was then attained by using previously identified lipids as internal standards. This, combined with an extremely high resolution (>800,000), yielded well-resolved isotopic fine structures allowing for the identification of lipids by MALDI 15-T FT-ICR MS without using tandem mass spectrometric (MS/MS) analysis. Using this method, a total of 38 unique lipids were successfully identified.

Two new triterpene caffeates have been isolated from the root bark of Hibiscus syriacus. Their structures were established through various spectral studies as 3beta,23,28-trihydroxy-12-oleanene 23-caffeate (1) and 3beta,23,28-trihydroxy-12-oleanene 3beta-caffeate (2). Compounds 1 and 2 showed lipid peroxidation inhibitory activity and significant cytotoxicity against a panel of human cancer cell lines.