Clamp loader ATPases and the evolution of DNA replication machinery

Clamp loaders are pentameric ATPases of the AAA+ family that operate to ensure processive DNA replication. They do so by loading onto DNA the ring-shaped sliding clamps that tether the polymerase to the DNA. Structural and biochemical analysis of clamp loaders has shown how, despite differences in composition across different branches of life, all clamp loaders undergo the same concerted conformational transformations, which generate a binding surface for the open clamp and an internal spiral chamber into which the DNA at the replication fork can slide, triggering ATP hydrolysis, release of the clamp loader, and closure of the clamp round the DNA. We review here the current understanding of the clamp loader mechanism and discuss the implications of the differences between clamp loaders from the different branches of life

Genetic variants in novel pathways influence blood pressure and cardiovascular disease risk.

Blood pressure is a heritable trait influenced by several biological pathways and responsive to environmental stimuli. Over one billion people worldwide have hypertension (≥140 mm Hg systolic blood pressure or  ≥90 mm Hg diastolic blood pressure). Even small increments in blood pressure are associated with an increased risk of cardiovascular events. This genome-wide association study of systolic and diastolic blood pressure, which used a multi-stage design in 200,000 individuals of European descent, identified sixteen novel loci: six of these loci contain genes previously known or suspected to regulate blood pressure (GUCY1A3-GUCY1B3, NPR3-C5orf23, ADM, FURIN-FES, GOSR2, GNAS-EDN3); the other ten provide new clues to blood pressure physiology. A genetic risk score based on 29 genome-wide significant variants was associated with hypertension, left ventricular wall thickness, stroke and coronary artery disease, but not kidney disease or kidney function. We also observed associations with blood pressure in East Asian, South Asian and African ancestry individuals. Our findings provide new insights into the genetics and biology of blood pressure, and suggest potential novel therapeutic pathways for cardiovascular disease prevention

Relatório de estágio em farmácia comunitária

Relatório de estágio realizado no âmbito do Mestrado Integrado em Ciências Farmacêuticas, apresentado à Faculdade de Farmácia da Universidade de Coimbr

A Computational Model of the LGI1 Protein Suggests a Common Binding Site for ADAM Proteins

Mutations of human leucine-rich glioma inactivated (LGI1) gene encoding the epitempin protein cause autosomal dominant temporal lateral epilepsy (ADTLE), a rare familial partial epileptic syndrome. The LGI1 gene seems to have a role on the transmission of neuronal messages but the exact molecular mechanism remains unclear. In contrast to other genes involved in epileptic disorders, epitempin shows no homology with known ion channel genes but contains two domains, composed of repeated structural units, known to mediate protein-protein interactions

Dynamics of open DNA sliding clamps

A range of enzymes in DNA replication and repair bind to DNA-clamps: torus-shaped proteins that encircle double-stranded DNA and act as mobile tethers. Clamps from viruses (such as gp45 from the T4 bacteriophage) and eukaryotes (PCNAs) are homotrimers, each protomer containing two repeats of the DNA-clamp motif, while bacterial clamps (pol III β) are homodimers, each protomer containing three DNA-clamp motifs. Clamps need to be flexible enough to allow opening and loading onto primed DNA by clamp loader complexes. Equilibrium and steered molecular dynamics simulations have been used to study DNA-clamp conformation in open and closed forms. The E. coli and PCNA clamps appear to prefer closed, planar conformations. Remarkably, gp45 appears to prefer an open right-handed spiral conformation in solution, in agreement with previously reported biophysical data. The structural preferences of DNA clamps in solution have implications for understanding the duty cycle of clamp-loaders

Defining the phenotypical spectrum associated with variants in TUBB2A

BACKGROUND: Variants in genes belonging to the tubulin superfamily account for a heterogeneous spectrum of brain malformations referred to as tubulinopathies. Variants in TUBB2A have been reported in 10 patients with a broad spectrum of brain imaging features, ranging from a normal cortex to polymicrogyria, while one patient has been reported with progressive atrophy of the cerebellar vermis. METHODS: In order to further refine the phenotypical spectrum associated with TUBB2A, clinical and imaging features of 12 patients with pathogenic TUBB2A variants, recruited via the international network of the authors, were reviewed. RESULTS: We report 12 patients with eight novel and one recurrent variants spread throughout the TUBB2A gene but encoding for amino acids clustering at the protein surface. Eleven patients (91.7%) developed seizures in early life. All patients suffered from intellectual disability, and 11 patients had severe motor developmental delay, with 4 patients (36.4 %) being non-ambulatory. The cerebral cortex was normal in five individuals and showed dysgyria of variable severity in seven patients. Associated brain malformations were less frequent in TUBB2A patients compared with other tubulinopathies. None of the patients had progressive cerebellar atrophy. CONCLUSION: The imaging phenotype associated with pathogenic variants in TUBB2A is highly variable, ranging from a normal cortex to extensive dysgyria with associated brain malformations. For recurrent variants, no clear genotype-phenotype correlations could be established, suggesting the role of additional modifiers

Retrospective evaluation of the dose equivalence of Botox and Dysport in the management of blepharospasm and hemifacial spasm: a novel paradigm for a never ending story

Botox(\uae) and Dysport(\uae) are the preparations of botulinum neurotoxin most widely used for therapeutic purposes. Several studies have addressed the topic of the equivalency ratio (D/B ratio) to be used in clinical practice and whether a reliable value exists is still a matter of debate. To this purpose, we ideated a novel paradigm by retrospectively examining the patients affected by hemifacial spasm and blepharospasm. We compared the pairs of treatments with a switch from one brand to the other undergone by the same patient in consecutive sessions with overlapping clinical outcome. Out of 2006 treatments, we found 51 treatment pairs. D/B ratio was extremely variable (range 1.2-13.3) and in most cases (65%) it was between 1:3 and 1:5. In conclusion, even if the 1:4 ratio might be reliable for clinical purpose, a true bioequivalence between Dysport(\uae) and Botox(\uae) might not exist due to the intrinsic difference in their pharmacokinetic properties