RESUMEN
Intrinsically disordered proteins (IDPs) lack stable tertiary structure under physiological conditions. The unique composition and complex dynamical behaviour of IDPs make them a challenge for structural biology and molecular evolution studies. Using NMR ensembles, we found that IDPs evolve under a strong site-specific evolutionary rate heterogeneity, mainly originated by different constraints derived from their inter-residue contacts. Evolutionary rate profiles correlate with the experimentally observed conformational diversity of the protein, allowing the description of different conformational patterns possibly related to their structure-function relationships. The correlation between evolutionary rates and contact information improves when structural information is taken not from any individual conformer or the whole ensemble, but from combining a limited number of conformers. Our results suggest that residue contacts in disordered regions constrain evolutionary rates to conserve the dynamic behaviour of the ensemble and that evolutionary rates can be used as a proxy for the conformational diversity of IDPs.
Asunto(s)
Proteínas Intrínsecamente Desordenadas/química , Modelos Moleculares , Conformación Proteica , Aminoácidos , Sitios de Unión , Evolución Molecular , Humanos , Proteínas Intrínsecamente Desordenadas/genética , Proteínas Intrínsecamente Desordenadas/metabolismo , Resonancia Magnética Nuclear Biomolecular , Unión Proteica , Relación Estructura-ActividadRESUMEN
Massive parallel sequencing technologies are facilitating the faster identification of sequence variants with the consequent capability of untangling the molecular bases of many human genetic syndromes. However, it is not always easy to understand the impact of novel variants, especially for missense changes, which can lead to a spectrum of phenotypes. This study presents a custom-designed multistep methodology to evaluate the impact of novel variants aggregated in the genome aggregation database for the HBB, HBA2, and HBA1 genes, by testing and improving its performance with a dataset of previously described alterations affecting those same genes. This approach scored high sensitivity and specificity values and showed an overall better performance than sequence-derived predictors, highlighting the importance of protein conformation and interaction specific analyses in curating variant databases. This study also describes the strengths and limitations of these structural studies and allows identifying residues in the globin chains more prone to tolerate substitutions.
Asunto(s)
Biología Computacional , Bases de Datos Genéticas , Variación Genética , Hemoglobinas/genética , Alelos , Sustitución de Aminoácidos , Biología Computacional/métodos , Biología Computacional/normas , Genotipo , Hemoglobinas/química , Humanos , Mutación con Pérdida de Función , Mutación , Sistemas de Lectura Abierta , Fenotipo , Sensibilidad y Especificidad , Globinas alfa/química , Globinas alfa/genética , Globinas beta/química , Globinas beta/genéticaRESUMEN
We describe here the molecular and hematological characteristics of novel frameshift mutations in exon 2 of the HBB gene (in heterozygous state) found in two Argentinean pediatric patients with dominant ß-thalassemia-like features. In Hb Wilde, HBB:c.270_273delTGAG(p.Glu90Cysfs*67), we detected the deletion of the third base of the codon 89 (T) and the codon 90 (GAG), whereas in Hb Patagonia, HBB:c.296_297dupGT(p.Asp99Trpfs*59), the frameshift mutation was due to a duplication of a 'GT' dinucleotide after the second base of codon 98 (GTG). The Hb Patagonia and Hb Wilde mutations would result in elongated ß-globin chains with modified C-terminal sequences and a total of 155 and 157 amino acids residues, respectively. Based on bioinformatics and structural analysis, as well as protein modeling, we predict that the elongated ß-globins would affect the formation of the αß dimers and their stability, which would further support the mechanism for the observed clinical features in both patients.
Asunto(s)
Variación Genética , Hemoglobinas Anormales/genética , Globinas beta/genética , Talasemia beta/diagnóstico , Talasemia beta/genética , Adolescente , Adulto , Recuento de Células Sanguíneas , Preescolar , Codón , Análisis Mutacional de ADN , Índices de Eritrocitos , Exones , Femenino , Mutación del Sistema de Lectura , Hemoglobinas Anormales/química , Humanos , Masculino , Modelos Moleculares , Polimorfismo de Nucleótido Simple , Conformación Proteica , Multimerización de Proteína , Globinas beta/químicaRESUMEN
Glycogen and starch are the major energy storage compounds in most living organisms. The metabolic pathways leading to their synthesis involve the action of several enzymes, among which glycogen synthase (GS) or starch synthase (SS) catalyze the elongation of the alpha-1,4-glucan backbone. At least five SS isoforms were described in Arabidopsis thaliana; it has been reported that the isoform III (SSIII) has a regulatory function on the synthesis of transient plant starch. The catalytic C-terminal domain of A. thaliana SSIII (SSIII-CD) was cloned and expressed. SSIII-CD fully complements the production of glycogen by an Agrobacterium tumefaciens glycogen synthase null mutant, suggesting that this truncated isoform restores in vivo the novo synthesis of bacterial glycogen. In vitro studies revealed that recombinant SSIII-CD uses with more efficiency rabbit muscle glycogen than amylopectin as primer and display a high apparent affinity for ADP-Glc. Fold class assignment methods followed by homology modeling predict a high global similarity to A. tumefaciens GS showing a fully conservation of the ADP-binding residues. On the other hand, this comparison revealed important divergences of the polysaccharide binding domain between AtGS and SSIII-CD.