Biology subjects

Correia, M. S. P.

Publications and source records attributed to Correia, M. S. P..

3 recordsLinked to original sources

bioRxiv2025

Systematic structural and functional analysis of metal-binding sites in native proteomes

Metal-binding proteins (MBPs) are widespread and critical for many biological processes. Proteomics-based approaches have identified MBPs in various systems but typically cannot determine the location of metal-binding sites or shed light on the functional roles of metals. Thus, a systems-wide map of metal binding sites and an understanding of the consequences of metal binding for protein function are still lacking. Here, we used limited proteolysis-coupled mass spectrometry (LiP-MS) combined with chelator treatment to globally map metal-protein interactions and binding sites in native protein lysates. We applied the pipeline, named Metal-LiP, to Escherichia coli, identified 981 peptides on 297 proteins that exhibited dose responsiveness to chelator treatment, and quantified the relative metal-binding affinity. These proteins are strongly enriched for known or predicted MBPs, based on our novel compiled ground truth dataset of previously annotated (n=1013) and predicted (n=850) MBPs. Further, chelator-responsive peptides mapped at or near annotated metal-binding sites in 3D protein structures, showing that Metal-LiP can identify metal-binding sites, and provided novel experimental evidence for previously predicted sites. Notably, Metal-LiP identified 225 novel candidate MBPs and we validated five of these novel MBPs (GatZ, AhpC, GrxB, MsrC, TrhO) using inductively coupled plasma-mass spectrometry to detect metal binding in purified proteins. Finally, we used orthogonal approaches to probe the broad functional consequences of metal binding. Using a combination of Metal-LiP, thermal proteome profiling, size-exclusion chromatography coupled to mass spectrometry, and structural analyses, we identified cases where metal binding affects protein stability, modulates the assembly states of protein complexes, or is likely to affect enzymatic activity. We thereby propose functional roles for metal binding in 98 of our Metal-LiP hits and in 225 proteins in total. In summary, we describe an approach to identify bona fide MBPs with binding site-level resolution across the proteome. We have combined it with other methods to identify 424 candidate MBPs in the E coli proteome, to map known and novel candidate metal-binding sites for 297 of these proteins, and to suggest functional roles of metal binding.

systems biology↗

bioRxiv2022

A Universal Language for Finding Mass Spectrometry Data Patterns

Even though raw mass spectrometry data is information rich, the vast majority of the data is underutilized. The ability to interrogate these rich datasets is handicapped by the limited capability and flexibility of existing software. We introduce the Mass Spec Query Language (MassQL) that addresses these issues by enabling an expressive set of mass spectrometry patterns to be queried directly from raw data. MassQL is an open-source mass spectrometry query language for flexible and mass spectrometer manufacturer-independent mining of MS data. We envision the flexibility, scalability, and ease of use of MassQL will empower the mass spectrometry community to take fuller advantage of their mass spectrometry data and accelerate discoveries.

bioinformatics↗

bioRxiv2021

Enterotoxigenic Escherichia coli display a distinct growth phase before entry into stationary phase with shifts in tryptophan- fucose- and putrescine metabolism and degradation of neurotransmitter precursors

Enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrhea in children and adults in endemic areas. Gene regulation of ETEC during growth in vitro and in vivo needs to be further evaluated, and here we describe the full transcriptome and metabolome of ETEC during growth from mid-logarithmic growth to stationary phase in rich medium (LB medium). We identified specific genes and pathways subjected to rapid transient alterations in gene expression and metabolite production during the transition between logarithmic to stationary growth. The transient phase during late exponential growth is different from the subsequent induction of stationary phase-induced genes, including stress and survival responses as described earlier. The transient phase was characterized by the repression of genes and metabolites involved in organic substance transport. Genes involved in fucose and putrescine metabolism were upregulated, and genes involved in iron transport were repressed. Expression of toxins and colonization factors were not changed, suggesting retained virulence. Metabolomic analyses showed that the transient phase was characterized by a drop of intracellular amino acids, e.g., L-tyrosine, L-tryptophan, L-phenylalanine, L-leucine, and L-glutamic acid, followed by increased levels at induction of stationary phase. A pathway enrichment analysis of the entire transcriptome and metabolome showed activation of pathways involved in the degradation of neurotransmitters aminobutyrate (GABA) and precursors of 5-hydroxytryptamine (serotonin). This work provides a comprehensive framework for further studies on transcriptional and metabolic regulation in pathogenic E. coli. ImportanceWe show that E. coli, exemplified by the pathogenic subspecies enterotoxigenic E. coli (ETEC), undergoes a stepwise transcriptional and metabolic transition into the stationary phase. At a specific entry point, E. coli induces activation and repression of specific pathways. This leads to a rapid decrease of intracellular levels of L-tyrosine, L-tryptophan, L-phenylalanine, L-leucine, and L-glutamic acid due to metabolism into secondary compounds. The resulting metabolic activity leads to an intense but short peak of indole production, suggesting that this is the previously described "indole peak," rapid decrease of intermediate molecules of bacterial neurotransmitters, increased putrescine and fucose uptake, increased glutathione levels, and decreased iron uptake. This specific transient shift in gene expression and metabolomics is short-lived and disappears when bacteria enter the stationary phase. We suggest it mainly prepares bacteria for ceased growth, but the pathways involved suggest that this transient phase substantially influences survival and virulence.

microbiology↗