We have determined the 2 A X-ray structure of the 219-residue N-terminal VHS and FYVE tandem domain unit of Drosophila Hrs. The unit assumes a pyramidal structure in which the much larger VHS domain (residues 1-153) forms a rectangular base and the FYVE domain occupies the apical end. The VHS domain is comprised of an unusual "superhelix" of eight alpha helices, and the FYVE domain is mainly built of loops, two double-stranded antiparallel sheets, and a helix stabilized by two tetrahedrally coordinated zinc atoms. The two-domain structure forms an exact 2-fold-related homodimer through antiparallel association of mainly FYVE domains. Dimerization creates two identical pockets designed for binding ligands with multiple negative charges such as citrate or phosphatidylinositol 3-phosphate.

The organophosphate acid anhydrolase (OPAA) is a member of a class of bimetalloenzymes that hydrolyze a variety of toxic acetylcholinesterase-inhibiting organophosphorus compounds, including fluorine-containing chemical nerve agents. It also belongs to a family of prolidases, with significant activity against various Xaa-Pro dipeptides. Here we report the X-ray structure determination of the native OPAA (58 kDa mass) from Alteromonas sp. strain JD6.5 and its cocrystal with the inhibitor mipafox [N,N'-diisopropyldiamidofluorophosphate (DDFP)], a close analogue of the nerve agent organophosphate substrate diisopropyl fluorophosphate (DFP). The OPAA structure is composed of two domains, amino and carboxy domains, with the latter exhibiting a "pita bread" architecture and harboring the active site with the binuclear Mn(2+) ions. The native OPAA structure revealed unexpectedly the presence of a well-defined nonproteinaceous density in the active site whose identity could not be definitively established but is suggestive of a bound glycolate, which is isosteric with a glycine (Xaa) product. All three glycolate oxygens coordinate the two Mn(2+) atoms. DDFP or more likely its hydrolysis product, N,N'-diisopropyldiamidophosphate (DDP), is present in the cocrystal structure and bound by coordinating the binuclear metals and forming hydrogen bonds and nonpolar interactions with active site residues. An unusual common feature of the binding of the two ligands is the involvement of only one oxygen atom of the glycolate carboxylate and the product DDP tetrahedral phosphate in bridging the two Mn(2+) ions. Both structures provide new understanding of ligand recognition and the prolidase and organophosphorus hydrolase catalytic activities of OPAA.

The family of about 50 periplasmic binding proteins, which exhibit diverse specificity (e.g., carbohydrates, amino acids, dipeptides, oligopeptides, oxyanions, metals, and vitamins) and range in size from 20 to 58 kDa, is a gold mine for an atomic-level investigation of structure and molecular recognition. These proteins serve as initial receptors for active transport systems or permeases. About six of these proteins, including the dipeptide-binding protein (DppA), are also primary receptors for chemotaxis. The structure of the unbound form of DppA (M(r) = 57,400) has been determined and refined to an R-factor of 0.169 to 2 A resolution. DppA consists of two distinct domains (I and II) connected by two "hinge" segments which form part of the base of the wide groove between the two domains. The relative orientation of the two domains gives the protein a pearlike shape, with domain I and domain II forming the larger and smaller apical ends, respectively. From the tip to the rounded bottom measures about 85 A, and the widest diameter is about 60 A. Domain I, which consists of two integrated subdomains, is folded from two separate polypeptide segments from the amino- and carboxyl-terminal ends. The more compact domain II is formed from the intervening segment. Comparison of the dipeptide-binding protein structure with that of the bound form of the similar oligopeptide-binding protein [Tame, J. R. H., Murshudov, G. N., Dodson, E. J., Neil, T. K., Dodson, G. G., Higgins, C. F., & Wilkinson, A. J. (1994) Science 264, 1578-1581] reveals the major features that differentiate the ligand specificity of the two proteins and describe the large hinge bending (about 55 degrees) between the two domains.

Neurotoxin-l from Nuju nuja oxiuna venom (NTX-1) has been crystallized by vapor diffusion in sitting drops. The crystals have cell dimensions of a = 25.2 A, b = 75.6 A, c = 35.9 A, and are in space group P2,2,2,. Three-dimensional data to 1.9A have been recorded by a Syntex P2, automatic diffractometer. The atomic structure of the toxin has been determined by molecular replacement using the a-cobratoxin (c&TX) as the search model. The position of 534 non-hydrogen protein atoms have been determined. The model contains 65 water molecules. Refinement has led to an R-factor of 19.3% at 1.9 A resolution. The secondary and tertiary structures of NTX-1 have been analyzed and a comparison with structure of the a-CTX has been made.Neurotoxin-1; X-ray structure; Nuju nuju oxiunu venom