Biology subjects

Pereira, H. D.

Publications and source records attributed to Pereira, H. D..

4 recordsLinked to original sources

bioRxiv2026

Experimental Validation of Coiled-Coil Architecture and Folding Dynamics in the Golgin Bug1

Golgins are widely described as long coiled-coil proteins that contribute to the structural organisation and trafficking functions of the Golgi apparatus. Although experimental structures have been determined for a limited number of golgin regions, atomic-level information on their extended coiled-coil segments remains scarce, and the oligomeric state, topology, and register of most predicted regions remain unestablished. Here, we characterise a predicted coiled-coil region of the yeast golgin Bug1 (BUG1cc) using structural, biophysical, and computational approaches. X-ray crystallography revealed a parallel, in-register dimeric coiled-coil containing ten heptad repeats and a predominantly hydrophobic core, with specific polar interactions contributing to dimer stabilisation. In solution, BUG1cc was dimeric under SEC-MALS conditions and remained highly alpha-helical across the pH and ionic strength conditions examined. CD measurements revealed pronounced scan-rate-dependent hysteresis, while DSC independently confirmed an asymmetry between heating and cooling transitions. Increasing protein concentration shifted both apparent transition temperatures while preserving thermal hysteresis, supporting chain association and conformational rearrangements in structural recovery. Structure-based simulations indicated that interface contacts and intra-chain helicity are thermodynamically coupled and melt as a single cooperative unit, and that the monomer released on dissociation is compact and only partially helical, so that reassociation proceeds through a coupled folding-binding mechanism whose rate-limiting step is conformational rather than bimolecular. Together, these results establish the molecular architecture of a predicted coiled-coil region of Bug1 and reveal a complex folding landscape in which oligomerisation, secondary-structure recovery, and kinetic barriers are tightly coupled.

biophysics↗

bioRxiv2020

Trypanosomatid selenophosphate synthetase structure, function and interaction with selenocysteine lyase

Early branching eukaryotes have been used as models to study the evolution of cellular molecular processes. Strikingly, human parasite of the Trypanosomatidae family (T. brucei, T. cruzi and L. major) conserve the complex machinery responsible for selenocysteine biosynthesis and incorporation in selenoproteins (SELENOK/SelK, SELENOT/SelT and SELENOTryp/SelTryp), although these proteins do not seem to be essential for parasite viability under laboratory controlled conditions. Selenophosphate synthetase (SEPHS/SPS) plays an indispensable role in selenium metabolism, being responsible for catalyzing the formation of selenophosphate, the biological selenium donor for selenocysteine synthesis. We solved the crystal structure of the L. major selenophosphate synthetase and confirmed that its dimeric organization is functionally important throughout the domains of life. We also demonstrated its interaction with selenocysteine lyase (SCLY) and showed that it is not present in other stable complexes involved in the selenocysteine pathway, namely the phosphoseryl-tRNASec kinase (PSTK)-Sec-tRNASec synthase (SEPSECS) and the tRNASec-specific elongation factor (eEFSec)-ribosome. Endoplasmic reticulum stress with ditiothreitol (DTT) or tunicamycin upon selenophosphate synthetase ablation in procyclic T. brucei cells led to a growth defect. On the other hand, only DTT presented a negative effect in bloodstream T. brucei expressing selenophosphate synthetase-RNAi. Although selenoprotein T (SELENOT) was dispensable for both forms of the parasite, SELENOT-RNAi procyclic T. brucei cells were sensitive to DTT. Together, our data suggest a role for the T. brucei selenophosphate synthetase in regulation of the parasites ER stress response. SynopsisSelenium is both a toxic compound and a micronutrient. As a micronutrient, it participates in the synthesis of specific proteins, selenoproteins, as the amino acid selenocysteine. The synthesis of selenocysteine is present in organisms ranging from bacteria to humans. The protozoa parasites of the Trypanosomatidae family, that cause major tropical diseases, conserve the complex machinery responsible for selenocysteine biosynthesis and incorporation in selenoproteins. However, this pathway has been considered dispensable for the protozoa cells. This has intrigued us, and lead to question that if maintained in the cell it should be under selective pressure and therefore be necessary. Also, since the intermediate products of selenocysteine synthesis are toxic to the cell, it has been proposed that these compounds need to be sequestered from the cytoplasm. Therefore, extensive and dynamic protein-protein interactions must happen to deliver those intermediates along the pathway. In this study we have investigated the molecular and structural interactions of different proteins involved in selenocystein synthesis and describe its involvement in the endoplasmic reticulum protection to oxidative stress. Our results also show how the interaction of different proteins leads to the protection of the cell against the toxic effects of seleium compounds during selenocysteine synthesis.

molecular biology↗

bioRxiv2017

Efficiency Of Genomic Prediction Of Non-Assessed Single Crosses

An important application of genomic selection in plant breeding is the prediction of untested single crosses (SCs). Most investigations on the prediction efficiency were based on tested SCs, using cross-validation. The main objective was to assess the prediction efficiency by correlating the predicted and true genotypic values of untested SCs (accuracy) and measuring the efficacy of identification of the best 300 untested SCs (coincidence), using simulated data. We assumed 10,000 SNPs, 400 QTLs, two groups of 70 selected DH lines, and 4,900 SCs. The heritabilities for the assessed SCs were 30, 60 and 100%. The scenarios included three sampling processes of DH lines, two sampling processes of SCs for testing, two SNP densities, DH lines from distinct and same populations, DH lines from populations with lower LD, two genetic models, three statistical models, and three statistical approaches. We derived a model for genomic prediction based on SNP average effects of substitution and dominance deviations. The prediction accuracy is not affected by the linkage phase. The prediction of untested SCs is very efficient. The accuracies and coincidences ranged from approximately 0.8 and 0.5, respectively, under low heritability, to 0.9 and 0.7, assuming high heritability. Additionally, we highlighted the relevance of the overall LD and evidenced that efficient prediction of untested SCs can be achieved for crops that show no heterotic pattern, for reduced training set size (10%), for SNP density of 1 cM, and for distinct sampling processes of DH lines, based on random choice of the SCs for testing.

genetics↗

bioRxiv2017

Efficiency of genome-wide association study in random cross populations

Genome-wide association studies (GWAS) with plant species have employed inbred lines panels. Our objectives were to present additional quantitative genetics theory for GWAS, evaluate the relative efficiency of GWAS in non-inbred and inbred populations and in an inbred lines panel, and assess factors affecting GWAS. Fifty samples of 400 individuals from populations with linkage disequilibrium were simulated. Individuals were genotyped for 10,000 single nucleotide polymorphisms (SNPs) and phenotyped for traits controlled by 10 quantitative trait loci (QTLs) and 90 minor genes, assuming different degrees of dominance and heritabilities of 40 and 80%. The average SNP density was 0.1 centiMorgan and the QTL heritabilities ranged from 3.2 to 11.8%. To increase the QTL detection power, the additive-dominance model must be fitted for traits controlled by dominance effects but must not be fitted for traits showing no dominance. The power of detection was maximized increasing the sample size to 400 and the false discovery rate (FDR) to 5%. The average power of detection for the low, intermediate, and high heritability QTLs were 9.7, 32.7, and 87.7%, respectively. Under sample size of 400 the observed FDR was equal to or lower than the specified level of significance. The association mapping was highly precise. The analysis of the inbred random cross population provided essentially the same results from the non-inbred population. The inbred lines panel provided the best results concerning the low and intermediate heritability QTL detection power, FDR, and mapping precision. The FDR is mainly affected by population structure, compared to relationship information.

genetics↗