RESUMO
Neutral lipases constitute one of the most ubiquitous and diverse families of enzymes. The recently solved crystal structures of three lipases show that enzymatic hydrolysis occurs with the assistance of a catalytic triad, which is structurally reminiscent of serine proteinases. However, these lipases only become active at the oil-water interface through a conformational change that exposes the active centre of the enzyme.
Assuntos
Lipase/química , Conformação Proteica , Sequência de Aminoácidos , Animais , Humanos , Dados de Sequência Molecular , Dobramento de Proteína , Relação Estrutura-AtividadeRESUMO
BACKGROUND: Many proteins undergo posttranslational modifications involving covalent attachment of lipid groups. Among them is palmitoylation, a dynamic, reversible process that affects trimeric G proteins and Ras and constitutes a regulatory mechanism for signal transduction pathways. Recently, an acylhydrolase previously identified as lysophospholipase has been shown to function as an acyl protein thioesterase, which catalyzes depalmitoylation of Galpha proteins as well as Ras. Its amino acid sequence suggested that the protein is evolutionarily related to neutral lipases and other thioesterases, but direct structural information was not available. RESULTS: We have solved the crystal structure of the human putative Galpha-regulatory protein acyl thioesterase (hAPT1) with a single data set collected from a crystal containing the wild-type protein. The phases were calculated to 1.8 A resolution based on anomalous scattering from Br(-) ions introduced in the cryoprotectant solution in which the crystal was soaked for 20 s. The model was refined against data extending to a resolution of 1.5 A to an R factor of 18.6%. The enzyme is a member of the ubiquitous alpha/beta hydrolase family, which includes other acylhydrolases such as the palmitoyl protein thioesterase (PPT1). CONCLUSIONS: The human APT1 is closely related to a previously described carboxylesterase from Pseudomonas fluorescens. The active site contains a catalytic triad of Ser-114, His-203, and Asp-169. Like carboxylesterase, hAPT1 appears to be dimeric, although the mutual disposition of molecules in the two dimers differs. Unlike carboxylesterase, the substrate binding pocket and the active site of hAPT1 are occluded by the dimer interface, suggesting that the enzyme must dissociate upon interaction with substrate.
Assuntos
Tioléster Hidrolases/química , Acilação , Sequência de Aminoácidos , Proteínas de Bactérias/química , Sítios de Ligação , Domínio Catalítico , Cristalografia por Raios X , Dimerização , Evolução Molecular , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Hidrolases/classificação , Modelos Moleculares , Dados de Sequência Molecular , Ácido Palmítico/metabolismo , Conformação Proteica , Processamento de Proteína Pós-Traducional , Proteínas Recombinantes de Fusão/química , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade da Espécie , Relação Estrutura-Atividade , Tioléster Hidrolases/classificaçãoRESUMO
BACKGROUND: The multidomain PDZ-RhoGEF is one of many known guanine nucleotide exchange factors that upregulate Rho GTPases. PDZ-RhoGEF and related family members play a critical role in a molecular signaling pathway from heterotrimeric G protein-coupled receptors to Rho proteins. A approximately 200 residue RGS-like (RGSL) domain in PDZ-RhoGEF and its homologs is responsible for the direct association with Galpha12/13 proteins. To better understand structure-function relationships, we initiated crystallographic studies of the RGSL domain from human PDZ-RhoGEF. RESULTS: A recombinant construct of the RGSL domain was expressed in Escherichia coli and purified, but it did not crystallize. Alternative constructs were designed based on a novel strategy of targeting lysine and glutamic acid residues for mutagenesis to alanine. A triple-point mutant functionally identical to the wild-type protein was crystallized, and its structure was determined by the MAD method using Se-methionine (Se-Met) incorporation. A molecular model of the RGSL domain was refined at 2.2 A resolution, revealing an all-helical tertiary fold with the mutations located at intermolecular lattice contacts. CONCLUSIONS: The first nine helices adopt a fold similar to that observed for RGS proteins, although the sequence identity with other such known structures is below 20%. The last three helices are an integral extension of the RGS fold, packing tightly against helices 3 and 4 with multiple hydrophobic interactions. Comparison with RGS proteins suggests features that are likely relevant for interaction with G proteins. Finally, we conclude that the strategy used to produce crystals was beneficial and might be applicable to other proteins resistant to crystallization.
Assuntos
Proteínas rho de Ligação ao GTP/química , Proteínas rho de Ligação ao GTP/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Cristalografia por Raios X , Epitopos , Subunidade alfa Gi2 de Proteína de Ligação ao GTP , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/química , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/metabolismo , Proteínas de Fluorescência Verde , Humanos , Proteínas Luminescentes/química , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Conformação Proteica , Dobramento de Proteína , Proteínas Proto-Oncogênicas/química , Proteínas Proto-Oncogênicas/metabolismo , Proteínas RGS/química , Proteínas RGS/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Homologia de Sequência de Aminoácidos , Transdução de Sinais , Proteínas rho de Ligação ao GTP/genéticaRESUMO
BACKGROUND: Neutral lipases are ubiquitous and diverse enzymes. The molecular architecture of the structurally characterized lipases is similar, often despite a lack of detectable homology at the sequence level. Some of the microbial lipases are evolutionarily related to physiologically important mammalian enzymes. For example, limited sequence similarities were recently noted for the Streptomyces exfoliatus lipase (SeL) and two mammalian platelet-activating factor acetylhydrolases (PAF-AHs). The determination of the crystal structure of SeL allowed us to explore the structure-function relationships in this novel family of homologous hydrolases. RESULTS: The crystal structure of SeL was determined by multiple isomorphous replacement and refined using data to 1.9 A resolution. The molecule exhibits the canonical tertiary fold of an alpha/beta hydrolase. The putative nucleophilic residue, Ser131, is located within a nucleophilic elbow and is hydrogen bonded to His209, which in turn interacts with Asp177. These three residues create a triad that closely resembles the catalytic triads found in the active sites of other neutral lipases. The mainchain amides of Met132 and Phe63 are perfectly positioned to create an oxyanion hole. Unexpectedly, there are no secondary structure elements that could render the active site inaccessible to solvent, like the lids that are commonly found in neutral lipases. CONCLUSIONS: The crystal structure of SeL reinforces the notion that it is a homologue of the mammalian PAF-AHs. We have used the catalytic triad in SeL to model the active site of the PAF-AHs. Our model is consistent with the site-directed mutagenesis studies of plasma PAF-AH, which implicate Ser273, His351 and Asp296 in the active site. Our study therefore provides direct support for the hypothesis that the plasma and isoform II PAF-AHs are triad-containing alpha/beta hydrolases.
Assuntos
Fosfolipases A/química , Streptomyces/enzimologia , 1-Alquil-2-acetilglicerofosfocolina Esterase , Sequência de Aminoácidos , Sítios de Ligação/fisiologia , Cristalografia por Raios X , Proteínas Fúngicas/química , Ligação de Hidrogênio , Lipase/química , Modelos Moleculares , Dados de Sequência Molecular , Fator de Ativação de Plaquetas/fisiologia , Estrutura Secundária de Proteína , Alinhamento de Sequência , Relação Estrutura-AtividadeRESUMO
Platelet-activating factor acetylhydrolases (PAF-AHs, EC 3.1.1.47) constitute a unique subfamily of phospholipases A(2), specific for short acyl chains in the sn-2 position of the phospholipid. Their primary substrate is the platelet-activating factor, PAF, from which they cleave an acetyl moiety with concomitant release of lysoPAF. However, some acetylhydrolase will also hydrolyze other polar phospholipids with up to 6-carbons long acyl chains in the sn-2 position. PAF-acetylhydrolases are diverse enzymes, and the well-characterized isoforms are serine-dependent hydrolases, which do not require Ca(2+) for activity. Given the existence of two pools of PAF, intra- and extracellular, the acetylhydrolases can be divided into two subclasses: those found in the cytosol and enzymes secreted to blood plasma or other body fluids. Recent crystallographic studies shed new light on the complex structure-function relationships in PAF-AHs.
Assuntos
Fosfolipases A/química , Fosfolipases A/metabolismo , 1-Alquil-2-acetilglicerofosfocolina Esterase , Animais , Encéfalo/enzimologia , Humanos , Modelos Moleculares , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
The crystal and molecular structure of a triacylglyceride lipase (EC 3.1.1.3) from the fungus Rhizomucor miehei was analyzed using X-ray single crystal diffraction data to 1.9 A resolution. The structure was refined to an R-factor of 0.169 for all available data. The details of the molecular architecture and the crystal structure of the enzyme are described. A single polypeptide chain of 269 residues is folded into a rather unusual singly wound beta-sheet domain with predominantly parallel strands, connected by a variety of hairpins, loops and helical segments. All the loops are right-handed, creating an uncommon situation in which the central sheet is asymmetric in that all the connecting fragments are located on one side of the sheet. A single N-terminal alpha-helix provides the support for the other, distal, side of the sheet. Three disulfide bonds (residues 29-268, 40-43, 235-244) stabilize the molecule. There are four cis peptide bonds, all of which precede proline residues. In all, 230 ordered water molecules have been identified; 12 of them have a distinct internal character. The catalytic center of the enzyme is made up of a constellation of three residues (His257, Asp203 and Ser144) similar in structure and function to the analogous (but not homologous) triad found in both of the known families of serine proteinases. The fourth residue in this system equivalent to Thr/Ser in proteinases), hydrogen bonded to Asp, is Tyr260. The catalytic site is concealed under a short amphipatic helix (residues 85 to 91), which acts as "lid", opening the active site when the enzyme is adsorbed at the oil-water interface. In the native enzyme the "lid" is held in place by hydrophobic interactions.
Assuntos
Proteínas Fúngicas/química , Lipase/química , Mucorales/enzimologia , Sequência de Aminoácidos , Catálise , Ligação de Hidrogênio , Lipase/metabolismo , Modelos Moleculares , Dados de Sequência Molecular , Conformação Proteica , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Difração de Raios XRESUMO
Close interactions of the C-H...O type are known to occur in a variety of organic crystals, although it had been often argued that they do not represent true hydrogen bonds. During an extensive comparative study of all structurally characterized serine hydrolases containing an Asp(Glu)-His-Ser catalytic triad at their active centers (i.e. serine proteinases, lipases, acetylcholinesterase and a thioesterase), we have discovered that the C epsilon 1 atom of the active site histidine is invariably in a close contact with a carbonyl oxygen. The stereochemistry of these contacts suggests a cohesive, predominantly electrostatic interaction, fully consistent with the requirements imposed by the generally accepted definition of a hydrogen bond. A study of a sample of protein structures refined at high resolution revealed that similar hydrogen bonds involving (His) C epsilon 1-H are found in approximately 15% of non-active site histidine residues. The ubiquitous occurrence of this hitherto underestimated contact in the active sites of serine hydrolases suggests functional significance. We propose that the (His)C epsilon 1-H...O=C bond affects the charge distribution within the imidazolium ion so as to weaken the N epsilon 2-H bond, thereby facilitating general acid catalysis by the active site histidine during both the acylation and deacylation steps of hydrolysis.
Assuntos
Acetilcolinesterase/química , Histidina/química , Lipase/química , Serina Endopeptidases/química , Sequência de Aminoácidos , Sítios de Ligação , Carbono/química , Ligação de Hidrogênio , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Molecular , Oxigênio/químicaRESUMO
Hydrogen bonds are a major feature of protein structure. By a generally accepted definition, they occur whenever a proton is shared by two electronegative atoms. Hence, only hydrogens bonded to nitrogen and oxygen atoms are usually considered in analyses of protein hydrogen bond networks. However, X-ray and neutron diffraction studies have shown that crystals of various organic compounds exhibit close C-H...X contacts (where X is an electronegative atom, in most cases oxygen) which show all the stereochemical hallmarks of hydrogen bonds. In this work, we describe an analysis of short C-H...O interactions in a sample of known protein structures representing different categories of tertiary folds and refined at a resolution of at least 2 A. Although our analysis is based on the calculated coordinates of hydrogen atoms, its results are statistically significant: we find strong evidence that a large percentage of short C...O contacts constitute cohesive interactions. Moreover, the stereochemical study of C-H...O = C contacts, in which the orientation of free electron orbitals on the acceptor oxygen atom can be predicted, reveals that these interactions exhibit stereochemical features typical of hydrogen bonds. Among the hydrogen atoms involved in these contacts, the most common are those bonded to alpha carbon. This is consistent with the fact that these hydrogens are more acidic than others. We describe four different categories of C-H...O = C bonds. Those found between C alpha-H groups and main chain oxygens in adjacent strands of beta sheets are the most ubiquitous. Our results call for a revision of crystallographic restrained refinement programs which treat close carbon-oxygen contacts as purely repulsive; they may also have implications for the understanding of some enzymatic reaction mechanisms.
Assuntos
Ligação de Hidrogênio , Proteínas/química , Carbono/química , Oxigênio/química , Dobramento de Proteína , Estrutura Secundária de Proteína , Água/químicaRESUMO
X-ray quality single crystals of the Escherichia coli thioesterase II have been obtained. The protein used for crystallization was overexpressed in parent organism. The crystals are orthorhombic, space group C222(1) with axial lengths a = 99.0 A, b = 121.1 A, c = 166.6 A. A complete homotetramer (120,000 Da) of four polypeptide chains, each 286 residues long, occupies the asymmetric unit. The diffraction pattern extends to approximately 2.4 A resolution using CuK alpha radiation from a rotating anode source. The elucidation of the three-dimensional structure of this unusual bacterial thioesterase will provide basis for the analysis of its unique catalytic mechanism and substrate specificity.
Assuntos
Escherichia coli/enzimologia , Ácido Graxo Sintases/química , Tioléster Hidrolases/química , Cristalização , Difração de Raios XRESUMO
The origin of co-operativity in haemoglobin (Hb) resides in the reduced affinity of the T-state. T-state Hb crystals grown from polyethyleneglycol can be liganded without the molecule switching to the R high affinity state. X-ray analysis of T-state alpha-oxy Hb and T-state met Hb has identified the structural basis for reduced affinity. The nature of the chemical tension at the haem environment is different in the alpha and beta haems. There are small but definite structural changes associated with ligation in the T-state: these prove to be mostly in the same direction as the larger changes that occur in the T-->R transition.
Assuntos
Hemoglobinas/química , Metemoglobina/química , Oxiemoglobinas/química , Simulação por Computador , Hemoglobinas/metabolismo , Ligantes , Metemoglobina/metabolismo , Modelos Moleculares , Oxigênio/metabolismo , Oxiemoglobinas/metabolismo , Estrutura Terciária de Proteína , Sais , Temperatura , Difração de Raios XRESUMO
X-ray quality single crystals of an extracellular esterase from pathogenic Streptomyces scabies were obtained by the hanging drop method. The crystals are monoclinic (space group C2, a = 161.1 A, b = 51.2 A, c = 124.2 A, beta = 100.6 degrees) with two molecules related by a noncrystallographic dyad in the asymmetric unit, with a solvent content of approximately 64%. The diffraction pattern from fresh crystals extends beyond 2 A resolution using sealed tube CuK alpha radiation. The study has been initiated in order to elucidate the mechanism of this unusual non-serine-dependent esterase, and to gain better understanding of the molecular basis of the pathogenesis of the scab disease.
Assuntos
Esterases/química , Streptomyces/enzimologia , Cristalização , Processamento de Imagem Assistida por Computador , Conformação Proteica , Difração de Raios XRESUMO
We have obtained X-ray quality single crystals of Vibrio harveyi acyltransferase. The protein was obtained from V. harveyi by a gene mobilization expression system. The crystals are monoclinic (space group P2(1), a = 89.9 A, b = 83.6 A, c = 47.1 A, beta = 97.3 degrees) with two molecules related by a pronounced non-crystallographic dyad in the asymmetric unit, with a solvent content of approximately 50%. The diffraction pattern from fresh crystals extends beyond 2 A resolution using sealed tube CuK alpha radiation. The elucidation of the three-dimensional structure of this enzyme, believed to contain a proteinase-like catalytic triad, which resembles in many ways other eukaryotic fatty acid chain terminating enzymes, may have important consequences for our understanding of the molecular basis of the final stages of the synthesis of fatty acids.
Assuntos
Aciltransferases/química , Vibrio/enzimologia , Aciltransferases/biossíntese , Aciltransferases/isolamento & purificação , Processamento de Imagem Assistida por Computador , Difração de Raios XRESUMO
A crystal structure of a totally inactive insulin molecule has been determined. For this insulin molecule, the first without detectable activity to be characterized, the A and B-chains are linked by a peptide bond between A1 Gly and B29 Lys. The molecule has retained all its normal self-association properties and it can also accommodate the two different conformations designated T and R, as seen in 4Zn native pig insulin crystals. The hexamers of the crosslinked insulin molecule were crystallized using the 4Zn insulin recipe of Schlichtkrull. The structure has been crystallographically refined with data extending to 2 A using restrained least-square methods. Comparison of the B29-A1 peptide crosslink insulin and the 4Zn native insulin reveals close structural similarities with the native dimer. The analysis of the structure confirms the earlier hypothesis that insulin structures in crystals are not in an active conformation and that a separation of N-terminal A-chain and C-terminal B-chain is required for interaction with the insulin receptor.
Assuntos
Insulina/química , Sequência de Aminoácidos , Sítios de Ligação , Simulação por Computador , Reagentes de Ligações Cruzadas , Substâncias Macromoleculares , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Difração de Raios X/métodos , Zinco/metabolismoRESUMO
Calsequestrin is the major Ca2+ binding protein in the lumen of the sarcoplasmic reticulum membranes. Two X-ray quality crystal forms of canine cardiac calsequestrin were obtained by the hanging drop method using KCl as a precipitant. One form is monoclinic (space group P2(1), a = 73.4 A, b = 104.4 A, c = 60.2 A, beta = 120.4 degrees) with two molecules in the asymmetric unit and a solvent content of approximately 40%. The second form is trigonal (P3(1)21 or P3(2)21, a = b = 99.3 A, c = 89.8 A) with a single molecule in the asymmetric unit and 55% solvent content. Cross rotation function calculations show that despite the different space groups the packing of the molecules in both crystals is likely to be similar suggesting the existence of a stable dimer. The monoclinic crystals diffract beyond 3 A using a laboratory rotating anode source, while under the same conditions the trigonal crystals diffract only to approximately 4.5 A. This is the first report of successful preparation of X-ray quality crystals of a high capacity Ca2+ binding protein.
Assuntos
Calsequestrina/química , Miocárdio/metabolismo , Conformação Proteica , Animais , Calsequestrina/isolamento & purificação , Cristalização , Cristalografia por Raios X/métodos , Cães , Retículo Sarcoplasmático/metabolismoRESUMO
The crystal structure of the common house mite (Dermatophagoides sp.) Der p 2 allergen was solved at 2.15 A resolution using the MAD phasing technique, and refined to an R-factor of 0.209. The refined atomic model, which reveals an immunoglobulin-like tertiary fold, differs in important ways from the previously described NMR structure, because the two beta-sheets are significantly further apart and create an internal cavity, which is occupied by a hydrophobic ligand. This interaction is structurally reminiscent of the binding of a prenyl group by a regulatory protein, the Rho guanine nucleotide exchange inhibitor. The crystal structure suggests that binding of non-polar molecules may be essential to the physiological function of the Der p 2 protein.
Assuntos
Alérgenos/química , Sequência de Aminoácidos , Animais , Antígenos de Dermatophagoides , Cristalização , Dissulfetos/química , Poeira , Epitopos , Escherichia coli/genética , Glicoproteínas , Ligação de Hidrogênio , Imunoglobulinas/química , Ligantes , Metionina/química , Ácaros , Modelos Moleculares , Conformação Proteica , Dobramento de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Selênio/química , Água/químicaRESUMO
The assembly of the insulin hexamer brings the six B13 glutamate side-chains at the centre into close proximity. Their mutual repulsion is unfavourable and zinc co-ordination to B10 histidine is necessary to stabilize the well known zinc-containing hexamers. Since B13 is always a carboxylic acid in all known sequences of hexamer forming insulins, it is likely to be important in the hormone's biology. The mutation of B13 Glu-->Gln leads to a stable zinc-free hexamer with somewhat reduced potency. The structures of the zinc-free B13 Gln hexamer and the 2Zn B13 insulin hexamer have been determined by X-ray analysis and refined with 2.5 A and 2.0 A diffraction data, respectively. Comparisons show that in 2Zn B13 Gln insulin, the hexamer structure (T6) is very like that of the native hormone. On the other hand, the zinc-free hexamer assumes a quaternary structure (T3/R3) seen in the native 4Zn insulin hexamer, and normally associated only with high chloride ion concentrations in the medium. The crystal structures show the B13 Gln side-chains only contact water in contrast to the B13 glutamate in 2Zn insulin. The solvation of the B13 Gln may be associated with this residue favouring helix at B1 to B8. The low potency of the B13 Gln insulin also suggests the residue influences the hormone's conformation.
Assuntos
Insulina/química , Conformação Proteica , Animais , Cristalização , Glutamatos , Ácido Glutâmico , Glutamina , Humanos , Insulina/metabolismo , Substâncias Macromoleculares , Modelos Moleculares , Mutagênese Sítio-Dirigida , Mutação , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/ultraestrutura , Suínos , Difração de Raios X , Zinco/químicaRESUMO
The structures of carbonmonoxyhaemoglobins A and Cowtown (His146 beta----Leu) have been refined at 2.2 A (1 A = 0.1 nm) and 2.3 A resolution, respectively. The least squares fit to the Fe-C-O line makes an angle to the haem normal of about 6 degrees. The Fe-C-O group is bent from linearity by about 7 degrees. The porphyrins in the CO liganded haemoglobins are ruffled. This deformation of the haem and the distortion of the Fe-C-O group may explain the low CO affinity of haemoglobin. The electron density for the C-terminal residues is low but sufficient to distinguish the histidyl and leucyl residues clearly. The similarity between these two structures, apart from 146 beta, means that the reduced alkaline Bohr effect is due solely to the replacement of histidine by a leucine.
Assuntos
Monóxido de Carbono/metabolismo , Carboxihemoglobina/ultraestrutura , Heme/metabolismo , Hemoglobinas Anormais/ultraestrutura , Hemoglobinas/metabolismo , Cristalografia , Humanos , Relação Estrutura-AtividadeRESUMO
The malonyl coenzyme A-acyl carrier protein transacylase, a single polypeptide chain of 358 amino acid residues and a molecular mass of 32 kDa, is a key component of the fatty acid synthase multienzyme complex. The elucidation of its three-dimensional structure will help in the understanding of the molecular basis of the biosynthesis of fatty acids, as well as of polyketides and related biologically active molecules. Three X-ray-quality crystal forms of the Escherichia coli fabD gene product encoding for malonyl coenzyme A-acyl carrier protein transacylase have been obtained using the hanging-drop method and ammonium sulfate as precipitant. Two are tetragonal and each contains two molecules in the asymmetric unit (form I: space group P4(3(1))2(1)2 with a = b = 83.9 A, c = 166.5 A and form II: space group P4 with a = b = 132.64 A, c = 38.85 A), whereas the third form belongs to the hexagonal system and contains one molecule in the asymmetric unit (space group P6(1(5)) with a = b = 68.52 A, c = 117.71 A). In each case, the diffraction pattern extends to approximately 2.0 A resolution using CuK alpha radiation from a rotating anode source.
Assuntos
Aciltransferases/química , Proteína de Transporte de Acila S-Maloniltransferase , Sequência de Aminoácidos , Proteínas de Bactérias/química , Cristalografia por Raios X , Escherichia coli/enzimologia , Proteínas de Escherichia coli , Ácido Graxo Sintase Tipo II , Dados de Sequência MolecularRESUMO
We have obtained well-ordered single crystals of the flavoenzyme trypanothione reductase from Crithidia fasciculata. The crystals are tetragonal rods with unit cell dimensions a = 128.6 A, c = 92.5 A. The diffraction pattern corresponds to a primitive lattice. Laue class 4/m. Diffraction to better than 2.4 A has been recorded at the Daresbury Synchrotron. The accurate elucidation of the three-dimensional structure of this enzyme is required to support the rational design of compounds active against a variety of tropical diseases caused by trypanosomal parasites.