Control Systems and Ornamental Lighting. A Case Study: Illumination of the Facade of Santiago Hospital in Úbeda (Jaén)

We propose the analysis of the control systems associated with ornamental lighting applied to the architectural and monumental heritage from our teaching experience developed since 2017 at the University of Córdoba—Illumination Faculty of Philosophy of Córdoba, year 2017, and Center of Experimental Culture in the Plaza de la Corredera, year 2018—and the lighting project of the facade of the Hospital de Santiago de Úbeda, Jaén. The DMX system is the one which mostly fulfill the requirements of this kind of lighting. Its implementation is increasingly simple and is associated in many cases with the type of luminaire to be used and the particularity of differential wiring. This solution allows a personalized, creative, and dynamic control and the adaptation of the daily lighting to different events of singular character. It can be used as an instrument of economic revitalization of the tourist and commercial sectors, and of diffusion of the architectural and historical heritage values. The application of the Internet of Things (IoT) and Big Data could allow the creation of a predictive model that helps the spectral design of light sources and an objective tool that contributes to the confluence of art and science

Naturally-Occurring Rare Mutations Cause Mild to Catastrophic Effects in the Multifunctional and Cancer-Associated NQO1 Protein

The functional and pathological implications of the enormous genetic diversity of the human genome are mostly unknown, primarily due to our unability to predict pathogenicity in a high-throughput manner. In this work, we characterized the phenotypic consequences of eight naturally-occurring missense variants on the multifunctional and disease-associated NQO1 protein using biophysical and structural analyses on several protein traits. Mutations found in both exome-sequencing initiatives and in cancer cell lines cause mild to catastrophic e ects on NQO1 stability and function. Importantly, some mutations perturb functional features located structurally far from the mutated site. These e ects are well rationalized by considering the nature of the mutation, its location in protein structure and the local stability of its environment. Using a set of 22 experimentally characterized mutations in NQO1, we generated experimental scores for pathogenicity that correlate reasonably well with bioinformatic scores derived from a set of commonly used algorithms, although the latter fail to semiquantitatively predict the phenotypic alterations caused by a significant fraction of mutations individually. These results provide insight into the propagation of mutational e ects on multifunctional proteins, the implementation of in silico approaches for establishing genotype-phenotype correlations and the molecular determinants underlying loss-of-function in genetic diseases.ERDF/Spanish Ministry of Science, Innovation and Universities-State Research Agency RTI2018-096246-B-I00 SAF2015-69796Junta de Andalucía P18-RT-241

Serological Findings in a Child with Paroxysmal Cold Haemoglobinuria

PCH is a rare autoimmune hemolytic anemia (AIHA) but is one of the most common causes of AIAH in children. For the diagnosis, it is important to perform the appropriate methods of serological investigation and show the typical biphasic reaction. This is a case report of a child who presented with features of haemolysis and was diagnosed with PCH of this way

A Dynamic Core in Human NQO1 Controls the Functional and Stability Effects of Ligand Binding and Their Communication across the Enzyme Dimer

Human NAD(P)H:quinone oxidoreductase 1 (NQO1) is a multi-functional protein whose alteration is associated with cancer, Parkinson’s and Alzheimer´s diseases. NQO1 displays a remarkable functional chemistry, capable of binding different functional ligands that modulate its activity, stability and interaction with proteins and nucleic acids. Our understanding of this functional chemistry is limited by the difficulty of obtaining structural and dynamic information on many of these states. Herein, we have used hydrogen/deuterium exchange monitored by mass spectrometry (HDXMS) to investigate the structural dynamics of NQO1 in three ligation states: without ligands (NQO1apo), with FAD (NQO1holo) and with FAD and the inhibitor dicoumarol (NQO1dic). We show that NQO1apo has a minimally stable folded core holding the protein dimer, with FAD and dicoumarol binding sites populating binding non-competent conformations. Binding of FAD significantly decreases protein dynamics and stabilizes the FAD and dicoumarol binding sites as well as the monomer:monomer interface. Dicoumarol binding further stabilizes all three functional sites, a result not previously anticipated by available crystallographic models. Our work provides an experimental perspective into the communication of stability effects through the NQO1 dimer, which is valuable for understanding at the molecular level the effects of disease-associated variants, post-translational modifications and ligand binding cooperativity in NQO1

Caenorhabditis elegans AGXT-1 is a mitochondrial and temperature-adapted ortholog of peroxisomal human AGT1: New insights into between-species divergence in glyoxylate metabolism

In humans, glyoxylate is an intermediary product of metabolism, whose concentration is finely balanced. Mutations in peroxisomal alanine:glyoxylate aminotransferase (hAGT1) cause primary hyperoxaluria type 1 (PH1), which results in glyoxylate accumulation that is converted to toxic oxalate. In contrast, glyoxylate is used by the nematode Caenorhabditis elegans through a glyoxylate cycle to by-pass the decarboxylation steps of the tricarboxylic acid cycle and thus contributing to energy production and gluconeogenesis from stored lipids. To investigate the differences in glyoxylate metabolism between humans and C. elegans and to determine whether the nematode might be a suitable model for PH1, we have characterized here the predicted nematode ortholog of hAGT1 (AGXT-1) and compared its molecular properties with those of the human enzyme. Both enzymes form active PLP-dependent dimers with high specificity towards alanine and glyoxylate, and display similar three-dimensional structures. Interestingly, AGXT-1 shows 5-fold higher activity towards the alanine/glyoxylate pair than hAGT1. Thermal and chemical stability of AGXT-1 is lower than that of hAGT1, suggesting temperature-adaptation of the nematode enzyme linked to the lower optimal growth temperature of C. elegans. Remarkably, in vivo experiments demonstrate the mitochondrial localization of AGXT-1 in contrast to the peroxisomal compartmentalization of hAGT1. Our results support the view that the different glyoxylate metabolism in the nematode is associated with the divergent molecular properties and subcellular localization of the alanine:glyoxylate aminotransferase activity.This work was supported by the Spanish Ministry of Science and Innovation (CSD2009-00088, BIO2012-34937 and SAF2011-23933), the Junta de Andalucia (P11-CTS-7187), and the Oxalosis and Hyperoxaluria Foundation (OHF2012 to B.C.). A.L.P. acknowledges a Ramon y Cajal research contract (RyC2009-04147) from the Spanish Ministry of Science and Innovation and the University of Granada. N. M-T acknowledges a FPI predoctoral fellowship from the Spanish Ministry of Science and Innovation. A.C.C. and N.T. were supported by the grant IOS-1353845 from the National Science Foundation (NSF). N.T. acknowledges the Tetelman Fellowship for International Research on the Sciences awarded by Yale University.Peer Reviewe

Counterintuitive structural and functional effects due to naturally occurring mutations targeting the active site of the disease-associated NQO1 enzyme*

Our knowledge on the genetic diversity of the human genome is exponentially growing. However, our capacity to establish genotype–phenotype correlations on a large scale requires a combination of detailed experimental and computational work. This is a remarkable task in human proteins which are typically multifunctional and structurally complex. In addition, mutations often prevent the determination of mutant high-resolution structures by X-ray crystallography. We have characterized here the effects of five mutations in the active site of the disease-associated NQO1 protein, which are found either in cancer cell lines or in massive exome sequencing analysis in human population. Using a combination of H/D exchange, rapid-flow enzyme kinetics, binding energetics and conformational stability, we show that mutations in both sets may cause counterintuitive functional effects that are explained well by their effects on local stability regarding different functional features. Importantly, mutations predicted to be highly deleterious (even those affecting the same protein residue) may cause mild to catastrophic effects on protein function. These functional effects are not well explained by current predictive bioinformatic tools and evolutionary models that account for site conservation and physicochemical changes upon mutation. Our study also reinforces the notion that naturally occurring mutations not identified as disease-associated can be highly deleterious. Our approach, combining protein biophysics and structural biology tools, is readily accessible to broadly increase our understanding of genotype–phenotype correlations and to improve predictive computational tools aimed at distinguishing disease-prone against neutral missense variants in the human genome

Digenic inheritance in cystinuria mouse model

Cystinuria is an aminoaciduria caused by mutations in the genes that encode the two subunits of the amino acid transport system b0,+, responsible for the renal reabsorption of cystine and dibasic amino acids. The clinical symptoms of cystinuria relate to nephrolithiasis, due to the precipitation of cystine in urine. Mutations in SLC3A1, which codes for the heavy subunit rBAT, cause cystinuria type A, whereas mutations in SLC7A9, which encodes the light subunit b0,+AT, cause cystinuria type B. By crossing Slc3a1-/- with Slc7a9-/- mice we generated a type AB cystinuria mouse model to test digenic inheritance of cystinuria. The 9 genotypes obtained have been analyzed at early (2- and 5-months) and late stage (8-months) of the disease. Monitoring the lithiasic phenotype by X-ray, urine amino acid content analysis and protein expression studies have shown that double heterozygous mice (Slc7a9+/-Slc3a1+/-) present lower expression of system b0,+ and higher hyperexcretion of cystine than single heterozygotes (Slc7a9+/-Slc3a1+/+ and Slc7a9+/+Slc3a1+/-) and give rise to lithiasis in 4% of the mice, demonstrating that cystinuria has a digenic inheritance in this mouse model. Moreover in this study it has been demonstrated a genotype/phenotype correlation in type AB cystinuria mouse model providing new insights for further molecular and genetic studies of cystinuria patients

Molecular Recognition of PTS-1 Cargo Proteins by Pex5p: Implications for Protein Mistargeting in Primary Hyperoxaluria

Peroxisomal biogenesis and function critically depends on the import of cytosolic proteins carrying a PTS1 sequence into this organelle upon interaction with the peroxin Pex5p. Recent structural studies have provided important insights into the molecular recognition of cargo proteins by Pex5p. Peroxisomal import is a key feature in the pathogenesis of primary hyperoxaluria type 1 (PH1), where alanine:glyoxylate aminotransferase (AGT) undergoes mitochondrial mistargeting in about a third of patients. Here, we study the molecular recognition of PTS1 cargo proteins by Pex5p using oligopeptides and AGT variants bearing different natural PTS1 sequences, and employing an array of biophysical, computational and cell biology techniques. Changes in affinity for Pex5p (spanning over 3–4 orders of magnitude) reflect different thermodynamic signatures, but overall bury similar amounts of molecular surface. Structure/energetic analyses provide information on the contribution of ancillary regions and the conformational changes induced in Pex5p and the PTS1 cargo upon complex formation. Pex5p stability in vitro is enhanced upon cargo binding according to their binding affinities. Moreover, we provide evidence that the rational modulation of the AGT: Pex5p binding affinity might be useful tools to investigate mistargeting and misfolding in PH1 by pulling the folding equilibria towards the native and peroxisomal import competent state