Purification, crystallization and preliminary X-ray analysis of haemoglobin I from the armoured catfish Liposarcus anisitsi

Liposarcus anisitsi is an armoured catfish that presents accessorial air oxygenation through a modified stomach, which allows this species to survive in waters with very low oxygen content. Analysis of its haemolysate has shown the presence of four haemoglobins; this work focuses on the main component, haemoglobin I. It has been crystallized in two different forms and X-ray diffraction data have been collected to 2.77 and 2.86 Angstrom resolution using synchrotron radiation. Crystals were determined to belong to the space groups C2 and P2(1) and preliminary structural analysis revealed the presence of one tetramer in the asymmetric unit in both crystal forms. The structure was determined using a standard molecular-replacement technique.

CRYSTALLIZATION AND PRELIMINARY DIFFRACTION DATA OF BOTHROPSTOXIN-I ISOLATED FROM THE VENOM OF BOTHROPS-JARARACUSSU

The myotoxic Lys-49 phospholipase bothropstoxin I was crystallized, and X-ray diffraction data were collected to 3.5 Angstrom resolution. Preliminary analysis reveals the presence of four molecules in the asymmetric unit.

Crystallization of the secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus 39E.

The secondary alcohol dehydrogenase from the thermophile Thermoanaerobacter ethanolicus 39E has been crystallized at 40 degrees C by vapour difussion using polyethelene glycol as a precipitant. The orthorhombic crystals belong to the space group P 2(1)2(1)2 with cell constants of a=170.0 Angstrom, b=125.7 Angstrom and c=80.5 Angstrom. A native X-ray diffraction data set has been collected to 2.7 Angstrom resolution.

Isolation, characterization and biological activity of acidic phospholipase A(2) isoforms from Bothrops jararacussu snake venom

Acidic phospholipase A(2) (PLA(2)) isoforms in snake venoms, particularly those from Bothrops jararacussu, have not been characterized. This article reports the isolation and partial biochemical, functional and structural characterization of four acidic PLA(2)s (designated SIIISPIIA, SIIISPIIB, SIIISPIIIA and SIIISPIIIB) from this venom. The single chain purified proteins contained 122 amino acid residues and seven disulfide bonds with approximate molecular masses of 15 kDa and isoelectric points of 5.3. The respective N-terminal sequences were: SIIISPIIA-SLWQFGKMIDYVMGEEGAKS; SIIISPIIB-SLWQFGKMIFYTGKNEPVLS; SIIISPIIIA-SLWQFGKMILYVMGGEGVKQ and SIIISPIIIB-SLWQFGKMIFYEMTGEGVL. Crystals of the acidic protein SIIISPIIIB diffracted beyond 1.8 Angstrom resolution. These crystals are monoclinic with unit cell dimensions of a = 40.1 Angstrom, b = 54.2 Angstrom and c = 90.7 Angstrom. The crystal structure has been refined to a crystallographic residual of 16.1% (R-free = 22.9%). Specific catalytic activity (U/mg) of the isolated acidic PLA(2)s were SIIISPIIA = 290.3 U/mg; SIIISPIIB = 279.0 U/mg; SIIISPIIIA = 270.7 U/mg and SIIISPIIIB = 96.5 U/mg. Although their myotoxic activity was low, SIIISPIIA, SIIISPIIIB and SIIISPIIIA showed significant anticoagulant activity. However, there was no indirect hemolytic activity. SIIISPIIIB revealed no anticoagulant, but presented indirect hemolytic activity. With the exception of SIIISPIIIB, which inhibited platelet aggregation, all the others were capable of inducing time-independent edema. Chemical modification with 4-bromophenacyl bromide did not inhibit the induction of edema, but did suppress other activities. (C) 2003 Editions scientifiques et medicales Elsevier SAS. All rights reserved.

Crystallization and preliminary X-ray diffraction analysis of the lectin from Canavalia gladiata seeds

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Crystallization and X-ray diffraction data analysis of human deoxyhaemoglobin A(0) fully stripped of any anions

In this work, initial crystallographic studies of human haemoglobin (Hb) crystallized in isoionic and oxygen-free PEG solution are presented. Under these conditions, functional measurements of the O-2-linked binding of water molecules and release of protons have evidenced that Hb assumes an unforeseen new allosteric conformation. The determination of the high-resolution structure of the crystal of human deoxy-Hb fully stripped of anions may provide a structural explanation for the role of anions in the allosteric properties of Hb and, particularly, for the influence of chloride on the Bohr effect, the mechanism by which Hb oxygen affinity is regulated by pH. X-ray diffraction data were collected to 1.87 Angstrom resolution using a synchrotron-radiation source. Crystals belong to the space group P2(1)2(1)2 and preliminary analysis revealed the presence of one tetramer in the asymmetric unit. The structure is currently being refined using maximum-likelihood protocols.

Crystallization, preliminary X-ray analysis and molecular-replacement solution of the carboxy form of haemoglobin I from the fish Brycon cephalus

Haemoglobin, the 'honorary enzyme' [Brunori (1999), Trends Biochem. Sci. 24, 158-161], constitutes a prime prototype for allosteric models. Here, the crystallization and preliminary X-ray analysis of haemoglobin I from the South American fish Brycon cephalus are reported. X-ray diffraction data have been collected to 2.5 Angstrom resolution using synchrotron radiation (LNLS). Crystals were determined to belong to the space group P6(1)22 and preliminary structural analysis revealed the presence of one dimer (alpha beta) in the asymmetric unit. The structure was determined using standard molecular-replacement techniques.

Disulfide Biochemistry in 2-Cys Peroxiredoxin: Requirement of Glu50 and Arg146 for the Reduction of Yeast Tsa1 by Thioredoxin

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Crystallization and preliminary X-ray crystallographic analysis of importin-alpha from Neurospora crassa

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Structure of the polypeptide crotamine from the Brazilian rattlesnake Crotalus durissus terrificus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Crystallization and preliminary X-ray diffraction studies of an L-amino-acid oxidase from Lachesis muta venom

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Preliminary X-ray crystallographic studies of a Lys49-phospholipase A(2) homologue from Bothrops pirajai venom complexed with p-bromophenacyl bromide and alpha-tocopherol inhibitors

PrTX-I, a non-catalytic and myotoxic Lys49-PLA(2) from Bothrops pirajai venom has been crystallized alone and in complex with bromophenacyl bromide (BPB), alpha-tocopherol and alpha-tocopherol acetate inhibitors. These crystals have shown to diffract X-rays between 2.34 and 1.65 angstrom resolution. All complexes crystals are isomorphous and belong to the space group P2(1) whereas native PrTX-I crystals belong to the P3(1)21.

Insights into metal ion binding in phospholipases A(2): ultra high-resolution crystal structures of an acidic phospholipase A(2) in the Ca2+ free and bound states

The electrophile Ca2+ is an essential multifunctional co-factor in the phospholipase A(2) mediated hydrolysis of phospholipids. Crystal structures of an acidic phospholipase A(2) from the venom of Bothrops jararacussu have been determined both in the Ca2+ free and bound states at 0.97 and 1.60 angstrom resolutions, respectively. In the Ca2+ bound state, the Ca2+ ion is penta-coordinated by a distorted pyramidal cage of oxygen and nitrogen atoms that is significantly different to that observed in structures of other Group I/II phospholipases A(2). In the absence of Ca2+, a water molecule occupies the position of the Ca2+ ion and the side chain of Asp49 and the calcium-binding loop adopts a different conformation. (c) 2005 Elsevier SAS. All rights reserved.

Crystal Structure Determination of Mebendazole Form A Using High-Resolution Synchrotron X-Ray Powder Diffraction Data

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Crystal structure of propylthiouracil determined using high-resolution synchrotron X-ray powder diffraction

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)