Biology subjects

Mas, G.

Publications and source records attributed to Mas, G..

4 recordsLinked to original sources

bioRxiv2026

Large-scale exploration of protein space by automated NMR

Protein structures can now be predicted and designed at scale, yet experimental access to dynamics and conformational heterogeneity remains limited in throughput. This gap prevents a systematic understanding of how protein sequences encode motion and functional flexibility. Here, we establish a scalable experimental pipeline combining protein design, automated production, and nuclear magnetic resonance (NMR) spectroscopy to enable high-throughput characterization of protein structure and dynamics at atomic resolution. A single operator can produce and analyze hundreds of isotopically labeled proteins per week, with per-sample cost largely defined by DNA synthesis. To benchmark this approach, we experimentally characterized 384 de novo designed proteins spanning diverse regions of structure space. High-quality two-dimensional NMR spectra were obtained for 239 samples (62% of designs overall). NMR characterization confirmed that the designed proteins adopt their intended folds, and revealed unexpected local dynamics that are not captured by current computational models. Our approach establishes a foundation for data-driven modelling of sequence-structure-dynamics relationships and unlocks a new regime of statistical structural biology, where insight into protein biophysics is gained from experimental ensemble studies of suitably designed protein clusters.

biophysics↗

bioRxiv2023

Toxin-mediated depletion of nicotinamide dinucleotides drives persister formation in a human pathogen

Toxin-antitoxin (TA) systems are widespread in bacteria and are implicated in genome stability, virulence, phage defense and persistence. Although TA systems encompass a large variety of molecular activities and cellular targets, their physiological role and regulatory mechanisms are often unclear1,2. Here, we show that a RES domain TA system increases the survival of the human pathogen P. aeruginosa during antibiotic treatment by generating a subpopulation of highly drug-tolerant persisters. The NatT toxin is an NAD phosphorylase, which leads to strong depletion of NAD and NADP in a subpopulation of cells. Actively growing P. aeruginosa cells effectively compensate for toxin-mediated NAD deficiency by inducing the NAD salvage path-way. In contrast, under nutrient-limited conditions, NatT generates NAD-depleted cells that give rise to drug tolerant persisters during outgrowth. Structural and biochemical analyses of active and inactive NatR-NatT complexes reveal how changes in NatR-NatT interaction controls toxin activity and autoregulation. Finally, we show that the NAD precursor nicotinamide blocks NatT activity and eliminates persister formation, exposing powerful metabolic feedback control of toxin activity. The findings that patient isolates contain natT gain-of-function alleles and that NatT increases P. aeruginosa virulence, argue that NatT contributes to P. aeruginosa fitness during infections. These studies provide mechanistic insight into how a TA system promotes pathogen persistence by disrupting essential metabolic pathways during nutrient stress.

microbiology↗

bioRxiv2019

Gene regulatory architectures dissect the evolutionary dynamics of regulatory elements in humans and non-human primates

To gain insight into the evolution of the epigenetic regulation of gene expression in primates, we extensively profiled a new panel of human, chimpanzee, gorilla, orangutan, and macaque lymphoblastoid cell lines (LCLs), using ChIP-seq for five histone marks, ATAC-seq and RNA-seq, further complemented with WGS and WGBS. We annotated regulatory elements and integrated chromatin contact maps to define gene regulatory architectures, creating the largest catalog of regulatory elements in primates to date. We report that epigenetic conservation and its correlation with sequence conservation in primates depends on the activity state of the regulatory element. Our gene regulatory architectures reveal the coordination of different types of components and highlight the role of promoters and intragenic enhancers in the regulation of gene expression. We observed that most regulatory changes occur in weakly active intragenic enhancers. Remarkably, novel human-specific intragenic enhancers with weak activities are enriched in human-specific mutations. These elements appear in genes with signals of positive selection, tissue-specific expression and particular functional enrichments, suggesting that the regulatory evolution of these genes may have contributed to human adaptation.

evolutionary biology↗

bioRxiv2018

Integrated NMR and cryo-EM atomic-resolution structure determination of a half-megadalton enzyme complex

Atomic-resolution structure determination is the key requirement for understanding protein function. Cryo-EM and NMR spectroscopy both provide structural information, but currently cryo-EM does not routinely give access to atomic-level structural data, and, generally, NMR structure determination is restricted to small (<30 kDa) proteins. We introduce an integrated structure determination approach that simultaneously uses NMR and EM data to overcome the limits of each of these methods. The approach enabled determination of the high-resolution structure of the 468 kDa large dodecameric aminopeptidase TET2 to a precision and accuracy below 1 [A] by combining secondary-structure information obtained from near-complete magic-angle-spinning NMR assignments of the 39 kDa-large subunits, distance restraints from backbone amides and specifically labelled methyl groups, and a 4.1 [A] resolution EM map. The resulting structure exceeds current standards of NMR and EM structure determination in terms of molecular weight and precision. Importantly, the approach is successful even in cases where only medium-resolution (up to 8 [A]) cryo-EM data are available, thus paving avenues for the structure determination of challenging biological assemblies.

biophysics↗