Homology modelling has matured into an important technique in structural biology, significantly contributing to narrowing the gap between known protein sequences and experimentally determined structures. Fully automated workflows and servers simplify and streamline the homology modelling process, also allowing users without a specific computational expertise to generate reliable protein models and have easy access to modelling results, their visualization and interpretation. Here, we present an update to the SWISS-MODEL server, which pioneered the field of automated modelling 25 years ago and been continuously further developed. Recently, its functionality has been extended to the modelling of homo- and heteromeric complexes. Starting from the amino acid sequences of the interacting proteins, both the stoichiometry and the overall structure of the complex are inferred by homology modelling. Other major improvements include the implementation of a new modelling engine, ProMod3 and the introduction a new local model quality estimation method, QMEANDisCo. SWISS-MODEL is freely available at https://swissmodel.expasy.org.
Motivation Methods that estimate the quality of a 3D protein structure model in absence of an experimental reference structure are crucial to determine a model’s utility and potential applications. Single model methods assess individual models whereas consensus methods require an ensemble of models as input. In this work, we extend the single model composite score QMEAN that employs statistical potentials of mean force and agreement terms by introducing a consensus-based distance constraint (DisCo) score. Results DisCo exploits distance distributions from experimentally determined protein structures that are homologous to the model being assessed. Feed-forward neural networks are trained to adaptively weigh contributions by the multi-template DisCo score and classical single model QMEAN parameters. The result is the composite score QMEANDisCo, which combines the accuracy of consensus methods with the broad applicability of single model approaches. We also demonstrate that, despite being the de-facto standard for structure prediction benchmarking, CASP models are not the ideal data source to train predictive methods for model quality estimation. For performance assessment, QMEANDisCo is continuously benchmarked within the CAMEO project and participated in CASP13. For both, it ranks among the top performers and excels with low response times. Availability and implementation QMEANDisCo is available as web-server at https://swissmodel.expasy.org/qmean. The source code can be downloaded from https://git.scicore.unibas.ch/schwede/QMEAN. Supplementary information Supplementary data are available at Bioinformatics online.
Recombinantly produced proteins are indispensable tools for medical applications. Since the majority of them are glycoproteins, their N-glycosylation profiles are major determinants for their activity, structural properties and safety. For therapeutical applications, a glycosylation pattern adapted to product and treatment requirements is advantageous. Physcomitrium patens (Physcomitrella, moss) is able to perform highly homogeneous complex-type N-glycosylation. Additionally, it has been glyco-engineered to eliminate plant-specific sugar residues by knock-out of the β1,2-xylosyltransferase and α1,3-fucosyltransferase genes (Δxt/ft). Furthermore, Physcomitrella meets wide-ranging biopharmaceutical requirements such as GMP compliance, product safety, scalability and outstanding possibilities for precise genome engineering. However, all plants, in contrast to mammals, lack the capability to perform N-glycan sialylation. Since sialic acids are a common terminal modification on human N-glycans, the property to perform N-glycan sialylation is highly desired within the plant-based biopharmaceutical sector. In this study, we present the successful achievement of protein N-glycan sialylation in stably transformed Physcomitrella. The sialylation ability was achieved in a Δxt/ft moss line by stable expression of seven mammalian coding sequences combined with targeted organelle-specific localization of the encoded enzymes responsible for the generation of β1,4-galactosylated acceptor N-glycans as well as the synthesis, activation, transport and transfer of sialic acid. Production of free (Neu5Ac) and activated (CMP-Neu5Ac) sialic acid was proven. The glycosidic anchor for the attachment of terminal sialic acid was generated by the introduction of a chimeric human β1,4-galactosyltransferase gene under the simultaneous knock-out of the gene encoding the endogenous β1,3-galactosyltransferase. Functional complex-type N-glycan sialylation was confirmed via mass spectrometric analysis of a stably co-expressed recombinant human protein.
The SIB Swiss Institute of Bioinformatics (www.isb-sib.ch) provides world-class bioinformatics databases, software tools, services and training to the international life science community in academia and industry. These solutions allow life scientists to turn the exponentially growing amount of data into knowledge. Here, we provide an overview of SIB's resources and competence areas, with a strong focus on curated databases and SIB's most popular and widely used resources. In particular, SIB's Bioinformatics resource portal ExPASy features over 150 resources, including UniProtKB/Swiss-Prot, ENZYME, PROSITE, neXtProt, STRING, UniCarbKB, SugarBindDB, SwissRegulon, EPD, arrayMap, Bgee, SWISS-MODEL Repository, OMA, OrthoDB and other databases, which are briefly described in this article.
Recombinantly produced proteins are indispensable tools for medical applications. Since the majority of them are glycoproteins, their N-glycosylation profiles are major determinants for their activity, structural properties and safety. For therapeutical applications, a glycosylation pattern adapted to product and treatment requirements is advantageous. Physcomitrella (Physcomitrium patens, moss) is able to perform highly homogeneous complex-type N-glycosylation. Additionally, it has been glyco-engineered to eliminate plant-specific sugar residues by knock-out of the β1,2-xylosyltransferase and α1,3-fucosyltransferase genes (Δxt/ft). Furthermore, P. patens meets wide-ranging biopharmaceutical requirements such as GMP compliance, product safety, scalability and outstanding possibilities for precise genome engineering. However, all plants, in contrast to mammals, lack the capability to perform N-glycan sialylation. Since sialic acids are a common terminal modification on human N-glycans, the property to perform N-glycan sialylation is highly desired within the plant-based biopharmaceutical sector. In this study, we present the successful achievement of protein N-glycan sialylation in stably transformed P. patens. The sialylation ability was achieved in a Δxt/ft moss line by stable expression of six mammalian coding sequences combined with targeted organelle-specific localization of the encoded enzymes responsible for synthesis, activation, transport and transfer of sialic acid. Production of free and (CMP)-activated sialic acid was proven. The glycosidic anchor for the attachment of terminal sialic acid was generated by the introduction of a chimeric human β1,4-galactosyltransferase gene under the simultaneous knock-out of the gene encoding the endogenous β1,3-galactosyltransferase. Functional complex-type N-glycan sialylation was confirmed via mass spectrometric analysis of a stably co-expressed recombinant human protein.
Qualitative changes in gene expression after an autopolyploidization event, a pure duplication of the whole genome, might be relevant for a different regulation of molecular mechanisms between angiosperms growing in a life cycle with a dominant diploid sporophytic stage and the haploid-dominant bryophytes. Whereas angiosperms repair DNA double strand breaks (DSB) preferentially via non-homologous end joining (NHEJ), in bryophytes homologous recombination (HR) is the main DNA-DSB repair pathway facilitating the precise integration of foreign DNA into the genome via gene targeting (GT). Here, we studied the influence of ploidy on gene expression patterns and GT efficiency in the moss Physcomitrella using haploid plants and autodiploid plants, generated via an artificial duplication of the whole genome. Single cells (protoplasts) were transfected with a GT construct and material from different time-points after transfection was analysed by microarrays and SuperSAGE sequencing. In the SuperSAGE data, we detected 3.7% of the Physcomitrella genes as differentially expressed in response to the whole genome duplication event. Among the differentially expressed genes involved in DNA-DSB repair was an upregulated gene encoding the X-ray repair cross-complementing protein 4 (XRCC4), a key player in NHEJ. Analysing the GT efficiency, we observed that autodiploid plants were significantly GT suppressed (p<0.001) attaining only one third of the expected GT rates. Hence, an alteration of global transcript patterns, including genes related to DNA repair, in autodiploid Physcomitrella plants correlated with a drastic suppression of HR.
Summary The auxin efflux PIN‐FORMED (PIN) proteins are conserved in all land plants and important players in plant development. In the moss Physcomitrella (Physcomitrium patens), three canonical PINs (PpPINA–C) are expressed in the leafy shoot (gametophore). PpPINA and PpPINB show functional activity in vegetative growth and sporophyte development. Here, we examined the role of PpPINC in the life cycle of Physcomitrella. We established reporter and knockout lines for PpPINC and analysed vegetative and reproductive tissues using microscopy and transcriptomic sequencing of moss gametangia. PpPINC is expressed in immature leaves, mature gametangia and during sporophyte development. The sperm cells (spermatozoids) of pinC knockout mutants exhibit increased motility and an altered flagella phenotype. Furthermore, the pinC mutants have a higher portion of differentially expressed genes related to spermatogenesis, increased fertility and an increased abortion rate of premeiotic sporophytes. Here, we show that PpPINC is important for spermatogenesis and sporophyte retention. We propose an evolutionary conserved way of polar growth during early moss embryo development and sporophyte attachment to the gametophore while suggesting the mechanical function in sporophyte retention of a ring structure, the Lorch ring.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.