Biology subjects

Stocchi, N.

Publications and source records attributed to Stocchi, N..

4 recordsLinked to original sources

bioRxiv2023

De novo assembly of the black flounder genome. Why do pleuronectiformes have such a small genome size?

Black flounder (Paralichthys orbignyanus) is an economically important ma-rine fish with aquaculture potential in Argentina due to its market value. In this study, we sequenced the whole genome using an Illumina sequencing technology. We started with two independent libraries (from one female and one pool of females; each with 150 bp paired-end reads, a mean insert length of 350 bp, and >35 X-fold coverage). Each library was assembled separately using SOAPdenovo2 and the resulting contigs were scaffolded with SSPACE3 before gaps were filled with GapCloser. In vertebrates, including teleosts, the number of transposable elements (TEs) is related to genome size, but it remains unclear whether the size of introns and exons also plays a role. Therefore, the main objective of the present study was to test whether the small genome size of Pleuronectiformes is related to the size of their introns and exons. The assemblies re-sulted in a genome size of [~]538 Mbp (41.35% GC content, 0.11% undetermined bases). Analysis of the assemblies at the core genes level (subset of the 458 universally ex-pressed KOG families) revealed that more than 98% of core genes are present, with more than 78% of them having more than 50% coverage. This indicates a fairly complete and accurate genome at the coding sequence level. Prediction of genes based on statistical predictors (geneid) and sequence-based predictors (Exonerate, using a closely related species, Paralichthys olivaceus, as a reference) was performed. This revealed 25,231 protein-coding genes, 445 tRNAs, 3 rRNAs, and more than 1,500 non-coding RNAs of other types (including a complete set of spliceosomes and several types of snoRNA and miRNA). As a result, this study concluded that the reduced genome size of flounders is related to a reduction in transcript size, mainly through a reduction in exon number, but also through a reduction in large introns. Thus, both components seem to be involved in the strategy of genome reduction in Pleuronectiformes.

genomics↗

bioRxiv2023

Comprehensive characterization of the complex BAHD acyltransferase family from 218 land plants species: phylogenomic analysis and identification of specificity determinant positions

Chemodiversity is a fundamental trait acquired by plants during their lands colonization. This resulted from an evolutionary process leading to the increase in the number of homologues from a distinct set of protein superfamilies, many of them associated to the specialized metabolism, which allowed the expansion of the chemical space to cope with several environmental cues. BAHD acyltransferases are among these important superfamilies, catalyzing a reaction leading to the acylation of acceptor metabolites with Coenzyme A-activated donors. BAHD acyltransferases can use a wide variety of substrates and they often times display substrate permissiveness towards a wide variety of substrates. Together, these factors complicates the reliable identification and functional annotation of BAHD homologues, also due to the (relatively) limited amount of biochemical data on BAHD acyltransferases. In this work, we take a phylogenomics and computational approach to study the BAHD superfamily in land plants. Using a clustered training set with 27 proteomes, followed by the classification of additional 191 proteomes, we obtained a final BAHDome with 15607 homologues. The training set was clustered in 16 groups, that, together with the identification of cluster of orthologues from the complete BAHDome, were partially assigned functionally to different families guided by the characterized activities (in terms of metabolites used) present in each group. However, the function assignation was not direct in several cases, due to large sequence number, taxonomical distribution on the group, and high sequence variability intra-group. Finally, we used the different families (and functional subfamilies) identified to detect specificity determining positions (SDPs), that may account to explain the functional diversity by finding key positions in, for example, substrate interaction. However, only a handful of the SDPs identified in this work are linked to the substrate binding pocket. Together, these results allow to partially annotate functional BAHD families, which have evolved in a complex pattern of taxonomical and functional signals to allow interaction with multiple substrates, more likely associated to protein dynamics rather than direct substrate interaction.

genomics↗

bioRxiv2022

Seqrutinator: Non-Functional Homologue Sequence Scrutiny for the Generation of large Datatsets for Protein Superfamily Analysis

BackgroundIn recent years protein bioinformatics has resulted in many good algorithms for multiple sequence alignment (MSA) and phylogeny. Little attention has been paid to sequence selection whereas notably recently published complete proteomes often have many sequences that are partial or derive from pseudogenes. Not only do these sequences add noise to the MSA, phylogeny and other downstream computational analyses, they also instigate many errors in the processing of the MSAs and downstream analyses, including the phylogeny. ObjectiveThis work aims to provide and test an objective, automated but flexible pipeline for the scrutiny of sequence sets from large, complex, eukaryotic protein superfamilies. The pipeline should classify sequences with high precision and recall as either functional or non-functional. The pipeline should classify no or only a few SwissProt sequences as non-functional (high precision) and sequences from other related superfamilies as non-functional (high recall) and result in a demonstrably much improved MSA (high performance). ResultsSeqrutinator is a pipeline that consists of five modules written in Python3 that identify and remove sequences that are likely Non-Functional Homologues (NFH). Here we tested the pipeline using three complex plant superfamilies (BAHD, CYP and UGT) that act in specialized metabolism, using the complete proteomes of 16 plant species as input and SwissProt as a control. Only 1.94% of SwissProt sequences with wetlab evidence were identified as NFH and all sequences from other related superfamilies were removed. Most NFH sequences are partial but, interestingly, their removal results in highly improved MSAs. a few but significant sequences that instigate large gaps were found. The five modules show similar behaviour when applied to the 16 sequence sets of the three analysed superfamilies. Pipelines with different module orders result in similar classifications and, moreover, show that different modules often detect the same sequences. Conclusion and perspectiveSeqrutinator forms a consistent pipeline for sequence scrutiny that does result in sequence sets that generate high fidelity MSAs. Recovery analyses show the method has high precision and recall.

bioinformatics↗

bioRxiv2017

Substrate binding and specificity appear as major forces in the functional diversification of eqolisins.

BackgroundEqolisins are rare acid proteases found in archaea, bacteria and fungi. Certain fungi secrete acids as part of their lifestyle and interestingly these also have many eqolisin paralogs, up to nine paralogs have been recorded. This suggests functional redundancy and diversification, which was the subject of the research we performed and describe here.\n\nResultsWe identified eqolisin homologs by means of iterative HMMER analysis of the NR database. The identified sequences were scrutinized for which we defined novel hallmarks, identified by molecular dynamics simulations of mutants of highly conserved positions, using the structure of an eqolisin that was crystallized in the presence of a transition state inhibitor. Four conserved glycines were shown to be required for functionality. A substitution of W67F is shown to be accompanied by the L105W substitution. Molecular dynamics shows that the W67 binds to the substrate via a {pi}-{pi} stacking and a salt bridge, the latter being stronger in a virtual W67F/L105W double mutant of the resolved structure of Scytalido-carboxyl peptidase-B (PDB ID: 2IFW)). Additional likely fatal mutants are discussed.\n\nUpon sequence scrutiny we obtained a set of 233 sequences that in all likelihood lack false positives. This was used to reconstruct a Bayesian phylogenetic tree. We identified 14 putative specificity determining positions (SDPs) of which four are explained by mere structural explanations and nine seem to correspond to functional diversification related wit substrate binding ans specificity. A first sub-network of SDPs is related to substrate specificity whereas the second sub-network seems to affect the dynamics of three loops that are involved in substrate binding.\n\nHighlightsEqolisins are acid proteases found in prokaryotes and fungi only.\n\nThe recently co-evolved W67F-L105W substitutions promote substrate binding\n\nTwo Specificity Determining Networks, SDN1 and 2, were identified\n\nSDN1 has four Specificity Determining Positions involved in substrate specificity\n\nSDN2 has five Specificity Determining Positions involved in loop-substrate dynamics

bioinformatics↗