Asunto(s)
Proteínas Bacterianas/química , Modelos Moleculares , Proteínas Represoras/química , Thermotoga maritima/química , Secuencia de Aminoácidos , Proteínas Bacterianas/genética , Cristalografía por Rayos X , Datos de Secuencia Molecular , Proteínas Represoras/genética , Análisis de Secuencia de ADN , Thermotoga maritima/genéticaRESUMEN
Glutathione S-transferases (GSTs) comprise a diverse superfamily of enzymes found in organisms from all kingdoms of life. GSTs are involved in diverse processes, notably small-molecule biosynthesis or detoxification, and are frequently also used in protein engineering studies or as biotechnology tools. Here, we report the high-resolution X-ray structure of Atu5508 from the pathogenic soil bacterium Agrobacterium tumefaciens (atGST1). Through use of comparative sequence and structural analysis of the GST superfamily, we identified local sequence and structural signatures, which allowed us to distinguish between different GST classes. This approach enables GST classification based on structure, without requiring additional biochemical or immunological data. Consequently, analysis of the atGST1 crystal structure suggests a new GST class, distinct from previously characterized GSTs, which would make it an attractive target for further biochemical studies.
Asunto(s)
Agrobacterium tumefaciens/enzimología , Proteínas Bacterianas/química , Glutatión Transferasa/química , Agrobacterium tumefaciens/química , Agrobacterium tumefaciens/citología , Secuencia de Aminoácidos , Proteínas Bacterianas/clasificación , Cristalografía por Rayos X , Dimerización , Glutatión Transferasa/clasificación , Modelos Moleculares , Datos de Secuencia Molecular , Filogenia , Estructura Secundaria de ProteínaAsunto(s)
Proteínas Bacterianas/química , Thermotoga maritima/enzimología , Secuencia de Aminoácidos , Proteínas Bacterianas/aislamiento & purificación , Cristalografía por Rayos X , Modelos Moleculares , Datos de Secuencia Molecular , Pliegue de Proteína , Estructura Secundaria de Proteína , Proteínas de Saccharomyces cerevisiae/química , Thermotoga maritima/química , Proteína de Unión al GTP ran/químicaAsunto(s)
Fosfatasa Ácida/química , Clostridium acetobutylicum/enzimología , Fosfatasa Ácida/aislamiento & purificación , Secuencia de Aminoácidos , Sitios de Unión , Clostridium acetobutylicum/química , Cristalografía por Rayos X , Modelos Moleculares , Datos de Secuencia Molecular , Conformación Proteica , Pliegue de ProteínaAsunto(s)
Dioxigenasas/química , Secuencia de Aminoácidos , Animales , Sitios de Unión , Cristalización , Bases de Datos de Proteínas , Dioxigenasas/genética , Dioxigenasas/metabolismo , Metales/química , Ratones , Modelos Moleculares , Datos de Secuencia Molecular , Estructura Molecular , Unión Proteica , Estructura Secundaria de Proteína , Estructura Terciaria de ProteínaAsunto(s)
Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Ciclofilinas/química , Ciclofilinas/metabolismo , Isomerasa de Peptidilprolil/química , Isomerasa de Peptidilprolil/metabolismo , Thermotoga maritima/enzimología , Secuencia de Aminoácidos , Proteínas Bacterianas/genética , Sitios de Unión , Cristalografía por Rayos X , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Isomerasa de Peptidilprolil/clasificación , Polietilenglicoles , Pliegue de Proteína , Estructura Cuaternaria de Proteína , Estructura Terciaria de ProteínaAsunto(s)
Ligasas de Carbono-Nitrógeno con Glutamina como Donante de Amida-N/química , Ligasas de Carbono-Nitrógeno con Glutamina como Donante de Amida-N/metabolismo , Thermotoga maritima/enzimología , Secuencia de Aminoácidos , Sitios de Unión , Cristalografía por Rayos X , Modelos Moleculares , Datos de Secuencia Molecular , Estructura Cuaternaria de Proteína , Estructura Terciaria de Proteína , Homología Estructural de ProteínaRESUMEN
The production of large numbers of highly purified proteins for X-ray crystallography is a significant bottleneck in structural genomics. At the Joint Center for Structural Genomics (JCSG; http://www.jcsg.org), specific automated protein expression, purification, and analytical methods are being utilized to study the proteome of Thermotoga maritima. Anion exchange and size exclusion chromatography (SEC), intended for the production of highly purified proteins, have been automated and the procedures are described here in detail. Analytical SEC has been included as a standard quality control test. A biological unit (BU) is the macromolecule that has been proven or is presumed to be functional. Correct assignment of BUs from protein structures can be difficult. BU predictions obtained via the Protein Quaternary Structure file server (PQS; http://pqs.ebi.ac.uk/) were compared to SEC data for 16 representative T. maritima proteins whose structures were solved at the JCSG, revealing an inconsistency in five cases. Herein, we report that SEC can be used to validate or disprove PQS-derived oligomeric models. A substantial amount of associated SEC and structural data should enable us to use certain PQS parameters to gauge the accuracy of these computational models and to generally improve their predictions.
Asunto(s)
Proteínas Bacterianas/química , Proteínas Bacterianas/aislamiento & purificación , Biología Computacional/métodos , Cristalografía por Rayos X/métodos , Proteómica/métodos , Thermotoga maritima/genética , Cromatografía en Gel/métodos , Biología Computacional/instrumentación , Conformación ProteicaRESUMEN
Nicotinamide adenine dinucleotide (NAD) is an essential cofactor for cellular redox reactions and can act as an important substrate in numerous biological processes. As a result, nature has evolved multiple biosynthetic pathways to meet this high chemical demand. In Saccharomyces cerevisiae, the NAD salvage pathway relies on the activity of nicotinic acid phosphoribosyltransferase (NAPRTase), a member of the phosphoribosyltransferase (PRTase) superfamily. Here, we report the structure of a eukaryotic (yeast) NAPRTase at 1.75 A resolution (locus name: YOR209C, gene name: NPT1). The structure reveals a two-domain fold that resembles the architecture of quinolinic acid phosphoribosyltransferases (QAPRTases), but with completely different dispositions that provide evidence for structural heterogeneity among the Type II PRTases. The identification of a third domain in NAPRTases provides a structural basis and possible mechanism for the functional modulation of this family of enzymes by ATP.