Pulmonary Artery Pseudoaneurysm in COVID-19-Associated Pulmonary Mucormycosis: Case Series and Systematic Review of the Literature

Mycopathologia - Literature on COVID-19-associated pulmonary mucormycosis (CAPM) is sparse. Pulmonary artery pseudoaneurysm (PAP) is an uncommon complication of pulmonary mucormycosis (PM), and...     

Large-scale prediction of function shift in protein families with a focus on enzymatic function

Protein function shift can be predicted from sequence comparisons, either using positive selection signals or evolutionary rate estimation. None of the methods have been validated on large datasets, however. Here we investigate existing and novel methods for protein function shift prediction, and benchmark the accuracy against a large dataset of proteins with known enzymatic functions. Function change was predicted between subfamilies by identifying two kinds of sites in a multiple sequence alignment: Conservation-Shifting Sites (CSS), which are conserved in two subfamilies using two different amino acid types, and Rate-Shifting Sites (RSS), which have different evolutionary rates in two subfamilies. CSS were predicted by a new entropy-based method, and RSS using the Rate-Shift program. In principle, the more CSS and RSS between two subfamilies, the more likely a function shift between them. A test dataset was built by extracting subfamilies from Pfam with different EC numbers that belong to the same domain family. Subfamilies were generated automatically using a phylogenetic tree-based program, BETE. The dataset comprised 997 subfamily pairs with four or more members per subfamily. We observed a significant increase in CSS and RSS for subfamily comparisons with different EC numbers compared to cases with same EC numbers. The discrimination was better using RSS than CSS, and was more pronounced for larger families. Combining RSS and CSS by discriminant analysis improved classification accuracy to 71%. The method was applied to the Pfam database and the results are available at http://FunShift.cgb.ki.se. A closer examination of some superfamily comparisons showed that single EC numbers sometimes embody distinct functional classes. Hence, the measured accuracy of function shift is underestimated.     

MutL homologs in restriction-modification systems and the origin of eukaryotic MORC ATPases

   

The provenance and biochemical roles of eukaryotic MORC proteins have remained poorly understood since the discovery of their prototype MORC1, which is required for meiotic nuclear division in animals. The MORC family contains a combination of a gyrase, histidine kinase, and MutL (GHKL) and S5 domains that together constitute a catalytically active ATPase module. We identify the prokaryotic MORCs and establish that the MORC family belongs to a larger radiation of several families of GHKL proteins (paraMORCs) in prokaryotes. Using contextual information from conserved gene neighborhoods we show that these proteins primarily function in restriction-modification systems, in conjunction with diverse superfamily II DNA helicases and endonucleases. The common ancestor of these GHKL proteins, MutL and topoisomerase ATPase modules appears to have catalyzed structural reorganization of protein complexes and concomitant DNA-superstructure manipulations along with fused or standalone nuclease domains. Furthermore, contextual associations of the prokaryotic MORCs and their relatives suggest that their eukaryotic counterparts are likely to carry out chromatin remodeling by DNA superstructure manipulation in response to epigenetic signals such as histone and DNA methylation.     

Hepatitis C virus internal ribosome entry site-mediated translation is stimulated by specific interaction of independent regions of human La autoantigen

The human La autoantigen has been shown to interact with the internal ribosome entry site (IRES) of hepatitis C virus (HCV) in vitro. Using a yeast three-hybrid system, we demonstrated that, in addition to full-length La protein, both N- and C-terminal halves were able to interact with HCV IRES in vivo. The exogenous addition of purified full-length and truncated La proteins in rabbit reticulocyte lysate showed dose-dependent stimulation of HCV IRES-mediated translation. However, an additive effect was achieved adding the terminal halves together in the reaction, suggesting that both might play critical roles in achieving full stimulatory activity of the full-length La protein. Using computational analysis, three-dimensional structures of the RNA recognition motifs (RRM) of the La protein were independently modeled. Of the three putative RRMs, RRM2 was predicted to have a good binding pocket for the interaction with the HCV IRES around the GCAC motif near the initiator AUG and RRM3 binds perhaps in a different location. This observation was further investigated by the filter-binding and toe-printing assays. The results presented here strongly suggest that both the N- and C-terminal halves can interact independently with the HCV IRES and are involved in stimulating internal initiation of translation.     

A new family of polymerases related to superfamily A DNA polymerases and T7-like DNA-dependent RNA polymerases

   

Using sequence profile methods and structural comparisons we characterize a previously unknown family of nucleic acid polymerases in a group of mobile elements from genomes of diverse bacteria, an algal plastid and certain DNA viruses, including the recently reported Sputnik virus. Using contextual information from domain architectures and gene-neighborhoods we present evidence that they are likely to possess both primase and DNA polymerase activity, comparable to the previously reported prim-pol proteins. These newly identified polymerases help in defining the minimal functional core of superfamily A DNA polymerases and related RNA polymerases. Thus, they provide a framework to understand the emergence of both DNA and RNA polymerization activity in this class of enzymes. They also provide evidence that enigmatic DNA viruses, such as Sputnik, might have emerged from mobile elements coding these polymerases.     

SUPFAM—a database of potential protein superfamily relationships derived by comparing sequence-based and structure-based families: implications for structural genomics and function annotation in genomes

Members of a superfamily of proteins could result from divergent evolution of homologues with insignificant similarity in the amino acid sequences. A superfamily relationship is detected commonly after the three-dimensional structures of the proteins are determined using X-ray analysis or NMR. The SUPFAM database described here relates two homologous protein families in a multiple sequence alignment database of either known or unknown structure. The present release (1.1), which is the first version of the SUPFAM database, has been derived by analysing Pfam, which is one of the commonly used databases of multiple sequence alignments of homologous proteins. The first step in establishing SUPFAM is to relate Pfam families with the families in PALI, which is an alignment database of homologous proteins of known structure that is derived largely from SCOP. The second step involves relating Pfam families which could not be associated reliably with a protein superfamily of known structure. The profile matching procedure, IMPALA, has been used in these steps. The first step resulted in identification of 1280 Pfam families (out of 2697, i.e. 47%) which are related, either by close homologous connection to a SCOP family or by distant relationship to a SCOP family, potentially forming new superfamily connections. Using the profiles of 1417 Pfam families with apparently no structural information, an all-against-all comparison involving a sequence-profile match using IMPALA resulted in clustering of 67 homologous protein families of Pfam into 28 potential new superfamilies. Expansion of groups of related proteins of yet unknown structural information, as proposed in SUPFAM, should help in identifying ‘priority proteins’ for structure determination in structural genomics initiatives to expand the coverage of structural information in the protein sequence space. For example, we could assign 858 distinct Pfam domains in 2203 of the gene products in the genome of Mycobacterium tubercolosis. Fifty-one of these Pfam families of unknown structure could be clustered into 17 potentially new superfamilies forming good targets for structural genomics. SUPFAM database can be accessed at http://pauling.mbu.iisc.ernet.in/~supfam.     

Comparative genomics uncovers novel structural and functional features of the heterotrimeric GTPase signaling system

Though the heterotrimeric G-proteins signaling system is one of the best studied in eukaryotes, its provenance and its prevalence outside of model eukaryotes remains poorly understood. We utilized the wealth of sequence data from recently sequenced eukaryotic genomes to uncover robust G-protein signaling systems in several poorly studied eukaryotic lineages such as the parabasalids, heteroloboseans and stramenopiles. This indicated that the Gα subunit is likely to have separated from the ARF-like GTPases prior to the last eukaryotic common ancestor. We systematically identified the structure and sequence features associated with this divergence and found that most of the neomorphic positions in Gα form a ring of residues centered on the nucleotide binding site, several of which are likely to be critical for interactions with the RGS domain for its GAP function. We also present evidence that in some of the potentially early branching eukaryotic lineages, like Trichomonas, Gα is likely to function independently of the Gβγ subunits. We were able to identify previously unknown Gγ subunits in Naegleria, suggesting that the trimeric version was already present by the time of the divergence of the heteroloboseans from the remaining eukaryotes. Evolution of Gα subunits is dominated by several independent lineage-specific expansions (LSEs). In most of these cases there are concomitant, independent LSEs of RGS proteins along with an extraordinary diversification of their domain architectures. The diversity of RGS domains from Naegleria in particular, which has the largest complement of Gα and RGS proteins for any eukaryote, provides new insights into RGS function and evolution. We uncovered a new class of soluble ligand receptors of bacterial origin with RGS domains and an extraordinary diversity of membrane-linked, redox-associated, adhesion-dependent and small molecule-induced G-protein signaling networks that evolved in early-branching eukaryotes, independently of parallel systems in animals. Furthermore, this newly characterized diversity of RGS domains helps in defining their ancestral conserved interfaces with Gα and also those interfaces that are prone to extensive lineage-specific diversification and are thereby responsible for selectivity in Gα-RGS interactions. Several mushrooms show LSEs of Gαs but not of RGS proteins pointing to the probable differentiation of Gαs in conjunction with mating-type diversity. When combined with the characterization of the 7TM receptors (GPCRs), it becomes apparent that, through much of eukaryotic evolution, cells contained both 7TM receptors that acted as GEFs and those as GAPs (with C-terminal RGS domains) for Gαs. Only in some lineages like animals and stramenopiles the 7TM receptors were restricted to GEF only roles, probably due to selection imposed by the rate-constants of the Gαs that underwent lineage-specific expansion in them. In the alveolate lineage the 7TM receptors occur independently of heterotrimeric G-proteins, suggesting the prevalence of G-protein-independent signaling in these organisms.     

Prediction of function divergence in protein families using the substitution rate variation parameter alpha

Protein families typically embody a range of related functions and may thus be decomposed into subfamilies with, for example, distinct substrate specificities. Detection of functionally divergent subfamilies is possible by methods for recognizing branches of adaptive evolution in a gene tree. As the number of genome sequences is growing rapidly, it is highly desirable to automatically detect subfamily function divergence. To this end, we here introduce a method for large-scale prediction of function divergence within protein families. It is called the alpha shift measure (ASM) as it is based on detecting a shift in the shape parameter (alpha [alpha]) of the substitution rate gamma distribution. Four different methods for estimating alpha were investigated. We benchmarked the accuracy of ASM using function annotation from Enzyme Commission numbers within Pfam protein families divided into subfamilies by the automatic tree-based method BETE. In a test using 563 subfamily pairs in 162 families, ASM outperformed functional site-based methods using rate or conservation shifting (rate shift measure [RSM] and conservation shift measure [CSM]). The best results were obtained using the "GZ-Gamma" method for estimating alpha. By combining ASM with RSM and CSM using linear discriminant analysis, the prediction accuracy was further improved.