RESUMEN
Haemoglobins constitute a set of proteins with interesting structural and functional properties, especially when the two large animal groups reptiles and fishes are focused on. Here, the crystallization and preliminary X-ray analysis of haemoglobin-II from the South American fish matrinxã (Brycon cephalus) is reported. X-ray diffraction data have been collected to 3.0 A resolution using synchrotron radiation (LNLS). Crystals were determined to belong to space group P2(1) and preliminary structural analysis revealed the presence of two tetramers in the asymmetric unit. The structure was determined using the standard molecular-replacement technique.
Asunto(s)
Peces/metabolismo , Hemoglobinas/química , Animales , Cromatografía DEAE-Celulosa , Cristalización , Cristalografía por Rayos X , Bases de Datos de Proteínas , Proteínas de Peces/química , Hemoglobinas/aislamiento & purificación , Programas InformáticosRESUMEN
The molecular structure of human uropepsin, an aspartic proteinase from the urine produced in the form of pepsinogen A in the gastric mucosa, has been determined by molecular replacement using human pepsin as the search model. Crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 50.99, b = 75.56, c = 89.90 A. Crystallographic refinement led to an R factor of 0.161 at 2.45 A resolution. The positions of 2437 non-H protein atoms in 326 residues have been determined and the model contains 143 water molecules. The structure is bilobal, consisting of two predominantly beta-sheet lobes which, as observed in other aspartic proteinases, are related by a pseudo-twofold axis. A model of the uropepsin-pepstatin complex has been constructed based on the high-resolution crystal structure of pepsin complexed with pepstatin.
Asunto(s)
Endopeptidasas/química , Secuencia de Aminoácidos , Sitios de Unión , Catálisis , Cristalización , Cristalografía por Rayos X , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Pepstatinas/metabolismo , Conformación Proteica , Control de Calidad , Homología de Secuencia de Aminoácido , Especificidad por SustratoRESUMEN
The three-dimensional structure of human uropepsin complexed with pepstatin has been modelled using human pepsin as a template. Uropepsin is an aspartic proteinase from the urine, produced in the form of pepsinogen A in the gastric mucosa. The structure is bilobal, consisting of two predominantly beta-sheet lobes which, as observed in other aspartic proteinases, are related by a pseudo twofold axis. A structural comparison between binary complexes of pepsin:pepstatin and uropepsin:pepstatin is discussed.
Asunto(s)
Endopeptidasas/química , Modelos Moleculares , Pepstatinas/química , Sitios de Unión , Humanos , Enlace de Hidrógeno , Conformación Proteica , Control de Calidad , Especificidad por SustratoRESUMEN
Considerable interest is currently focused on fish haemoglobins in order to identify the structural basis for their diversity of functional behaviour. The armored catfish Liposarcus anisitsi presents accessorial air breathing through a modified stomach, which allows this species to survive in waters with low oxygen content. The analysis of its haemolysate has shown the presence of four main haemoglobins, with this work focusing on haemoglobin IV (LaHb-IV). LaHb-IV was crystallized and X-ray diffraction data were collected to 2.4 A resolution using a synchrotron-radiation source. The crystal belongs to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 52.6 (1), b = 104.8 (2), c = 113.9 (2) A; preliminary structural analysis revealed the presence of one tetramer in the asymmetric unit. The structure was determined using the standard molecular-replacement technique.
Asunto(s)
Hemoglobinas/química , Animales , Bagres , Cristalización , Cristalografía por Rayos X , Hemoglobinas/aislamiento & purificación , Modelos Moleculares , Conformación ProteicaRESUMEN
Mastoparans are tetradecapeptides found to be the major component of vespid venoms. These peptides present a wide spectrum of biological activities, such as mast cell degranulation, hemolytic activity and also reveals antimicrobial activity. A mastoparan toxin isolated from the venom of Anterhynchium flavomarginatum micado has been crystallized. At room temperature these crystals diffracted to 2.8 A resolution. However, upon cooling to cryogenic temperature around 85 K, the original resolution limit could be improved to 2.0 A. Crystals were determined to belong to the space group P3(1) (P3(2)). This is the first mastoparan to be crystallized and it will provide further insights in the conformational significance of mastoparan toxins, with respect to their potency and activity in G protein regulation.
Asunto(s)
Cristalografía por Rayos X/métodos , Proteínas de Insectos/química , Venenos de Avispas/química , Avispas/metabolismo , Animales , Frío , Cristalización , Proteínas de Insectos/aislamiento & purificación , Venenos de Avispas/aislamiento & purificaciónRESUMEN
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 A 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 (alphabeta) in the asymmetric unit. The structure was determined using standard molecular-replacement techniques.
Asunto(s)
Hemoglobinas Anormales/química , Hemoglobinas , Adenosina Trifosfato/metabolismo , Animales , Cristalización , Cristalografía por Rayos X , Peces , Hemoglobinas Anormales/aislamiento & purificación , Hemoglobinas Anormales/metabolismo , Conformación ProteicaRESUMEN
Mastoparans are tetradecapeptides found to be the major component of vespid venoms. A mastoparan toxin isolated from the venom of Anterhynchium flavomarginatum micado has been crystallized and X-ray diffraction data collected to 2.7 A resolution using a synchrotron-radiation source. Crystals were determined to belong to the space group P6(2)22 (P6(4)22). This is the first mastoparan to be crystallized and will provide further insights into the conformational significance of mastoparan toxins with respect to their potency and activity in G-protein regulation.
Asunto(s)
Degranulación de la Célula , Mastocitos/citología , Venenos de Avispas/química , Cristalización , Cristalografía por Rayos X , Conformación ProteicaRESUMEN
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 A 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.
Asunto(s)
Hemoglobinas Anormales/química , Hemoglobinas , Animales , Bagres , Cristalización , Cristalografía por Rayos X , Hemoglobinas Anormales/aislamiento & purificación , SincrotronesRESUMEN
Carboxyhaemoglobin-II isolated from the pacu (Piaractus mesopotamicus) has been crystallized and X-ray diffraction data were collected to 2.0 A resolution using synchrotron radiation. Crystals were characterized as belonging to the space group I23; preliminary structural analysis reveals the presence of one dimer in the asymmetric unit.
Asunto(s)
Carboxihemoglobina/química , Cipriniformes/sangre , Animales , Carboxihemoglobina/aislamiento & purificación , Cristalización , Cristalografía por Rayos X , Modelos Moleculares , Conformación ProteicaRESUMEN
Oxyhaemoglobin I isolated from the Brazilian wolf Chrysocyon brachiurus has been crystallized and X-ray diffraction data has been collected to 2.06 A resolution using a synchrotron-radiation source. Crystals were determined to belong to the space group P2(1)2(1)2(1) and preliminary structural analysis revealed the presence of one tetramer in the asymmetric unit. The structure was determined using standard molecular-replacement techniques and is currently being refined using maximum-likelihood protocols. This is the first haemoglobin isolated from a member of the Canidae family to be crystallized and it will provide further insights in the comparative biochemistry of vertebrate haemoglobins.