Biology subjects

Marti, J.

Publications and source records attributed to Marti, J..

16 recordsLinked to original sources

bioRxiv2026

Conformational drugging pockets at the surface of c-MYC proto-oncongenes

The MYC proto-oncogene encodes a master transcription factor essential for regulating cellular homeostasis, growth, and metabolism. However, its deregulation is involved in up to 70% of human cancers, where it drives uncontrolled cellular proliferation. Structurally, MYC is characterized by an N-terminal transactivation domain and a C-terminal bHLHLZ domain critical for DNA binding and dimerization with MAX. Despite its clinical significance, MYC has remained historically "undruggable" due to its intrinsically disordered structure and the absence of traditional binding pockets. In this study, we combined all-atom Molecular Dynamics and well-tempered Metadynamics simulations to map the conformational landscape of a MYC motif in aqueous ionic solution. The selected energy-minimised structures exhibit stable conformations and drug pockets, providing structural foundations for targetting c-MYC, which is traditionally considered "undruggable". Furthermore, both the electrostatic potential and docking results for the obtained structure and the MYC inhibitor 10074-G5 demonstrated excellent fit, highlighting its potential as a drug target. Remarkably, MYC maintained steady binding to 10074-G5 throughout the 500 ns simulation. These findings provide a previously uncharacterized structural foundation for the design of innovative therapeutic interventions, such as small-molecule inhibitors or peptides aimed at disrupting oncogenic MYC activity.

cancer biology↗

bioRxiv2026

Nucleotide-dependent Structural Selection Governs c-Src Phosphorylation of Oncogenic KRas4B-G12D

c-Src, the first identified oncogene, and KRas, one of the most frequently mutated proteins in human cancers, can activate each other to promote tumorigenesis. Although normal KRas proteins cycle between the inactive GDP bound and the active GTP bound forms, oncogenic mutants are predominantly GTP bound. Importantly, c-Src preferentially recognizes and phosphorylates the GTP-bound form of KRas but the molecular mechanism underlying this specific recognition remains unknown. Here, we employ molecular simulation tools to identify the mechanism underlying the c-Src recognition of the GTP-loaded state of the G12D mutant of KRas4B (the most prevalent in humans). Combining extensive all-atom Molecular Dynamics simulations and Markov State Models analysis, we found that the most populated states of GTP-bound KRas maintain more open and dynamic switch regions, facilitating easier access of c-Src to the phosphorylation sites of KRas (Tyr32 and Tyr64). These states are sparsely populated for GDP loaded KRas. Docking calculations refined by molecular dynamics identify two c-Src specific regions (residues 340-359 and 453-473) able to stabilize phosphorylation-competent KRas conformations. Therefore, c-Src engages highly populated macrostates of GTP-loaded KRas, while interactions with the GDP form are limited to rare conformations. These KRas conformations selectively recognized by c-Src constitute privileged targets for the rational design of peptide-based or small-molecule inhibitors that specifically target active KRas4B-G12D while sparing the inactive GDP-bound form.

biophysics↗

bioRxiv2025

Conformational Dynamics of the Free MAX Protein Revealed by Well-Tempered Metadynamics

One of most relevant challenges in tumorigenesis is the association of MYC and MAX proteins, whose related cancers remain undrugged. In particular, the disordered regions shown by those oncogenes make their structural characterization and the development of new drugs a truly hard task. To address these challenges, we employed the enhanced-sampling well-tempered metadynamics method to systematically explore the conformational space of the unbound MAX protein. Our results revealed, for the first time, a well-defined and thermodynamically favorable conformation of monomeric MAX. This is a remarkable finding, as it demonstrates that regions of MAX previously considered persistently disordered are capable of adopting stable, folded structures under specific conditions. Moreover, our findings also suggest that the meta-stable structural motifs observed in this work may harbor druggable sites, particularly relevant for strategies aiming to target MAX directly or to disrupt its interaction with MYC, thereby modulating oncogenic signaling pathways. The present study establishes a new structural framework for understanding the dynamics of MAX and provides a foundation for future structure-based drug design targeting the MYC/MAX axis. Finally, our work offers a strategic blueprint for investigating similarly challenging drug targets. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/673476v3_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@18ea239org.highwire.dtl.DTLVardef@13c8b8eorg.highwire.dtl.DTLVardef@1ee414corg.highwire.dtl.DTLVardef@10114c1_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2024

Quantitative comparison of the structural differences between NRAS and its mutations by well-tempered metadynamics simulations

The NRAS-mutant subset of melanoma is one of the most aggressive and lethal types associated with poor overall survival. Unfortunately, a low understanding of the NRAS-mutant dynamic behavior has lead to the lack of clinically approved therapeutic agents able to directly target NRAS oncogenes. In this work, accurate local structures of NRAS and its mutants have been fully explored through the corresponding free energy surfaces obtained by microsecond scale well-tempered metadynamics simulations. Free energy calculations are crucial to reveal the precise mechanisms of Q61 mutations at the atomic level. Considering specific atom-atom distances d and angles{phi} as appropriate reaction coordinates we have obtained free energy surfaces revealing local and global minima together with their main transitions states, unvealing the mechanisms of abnormal NRAS activation from atomic-level and quantitatively analyzing the corresponding stable states. This will help to advance in our understanding of the basic mechanisms of NRAS mutations, offering new opportunities for the design of potential inhibitors.

biophysics↗

bioRxiv2024

Free energy profiles of temozolomide crossing brain plasma membranes

Temozolomide is an efficient small-molecule drug mostly employed for the treatment of glioblastoma, a tumor attacking both the spinal cord and the brain. Understanding the interactions of temozolomide with different lipids at the brain cell membrane can help elucidate how temozolomide permeates through cell membranes and its membrane-crossing ability. In the present work, we have constructed a simplified brain plasma membrane model to explore temozolomides microscopic structure and dynamics by means of all-atom microsecond scale molecular dynamics simulations. The preferential location of temozolomide is at the solvent-aqueous fluid surrounding the brain membrane, but it can access the interface with the membrane regularly, eventually binding to lipids of the choline and cerebroside classes. The free energy barriers of temozolomide related to brain-like plasma membrane crossing were investigated by adaptive biasing force methods, revealing values ranging from 18.5 to 66.5 kcal/mol at temperatures of 323 K and 310 K, respectively. Our results suggest that temozolomide cannot cross the membrane by pure diffusion at the normal human body temperature but that rising the temperature significantly increases the probability of barrier crossing. This fact is mainly due to the crucial role played by cholesterol and lipids of the cerebroside class. The findings reported in this work can be used to optimize the molecular design of temozolomide and to develop new analogs with better pharmacokinetic properties. Author summaryGlioblastoma is a devastating tumor affecting the brain and spinal cord, which has in the FDA-approved drug temozolomide its main clinical treatment. The present study explores how temozolomide interacts with several lipids in brain-like cell membranes. Our findings show that at normal body temperature temozolomide cannot cross the membrane by pure diffusion, but that higher temperatures significantly enhance its ability to cross the membrane by reducing the free energy barriers. Temozolomide interacts differently with several lipids and sterols depending on the temperature, which affects its permeability. This implies that temozolomide will cross the outer layer of the brain membrane only with the help of driving agents, such as intermembrane proteins. Our research suggests that temozolomide may be more effective at higher temperatures and cancer patients with fever might need a lower dose. Importantly, cholesterol plays a key role in blocking temozolomide from crossing brain-like membranes, so reducing dietary intake of cholesterol and cerebroside lipids could help modify brain cell membranes, making it easier for temozolomide to target cancer cells effectively and potentially reducing side effects.

biophysics↗

bioRxiv2024

Atomic-scale mechanisms of GDP extraction by SOS1 in KRAS-G12 and KRAS-D12 oncogenes

The guanine exchange factor SOS1 is a crucial node into the positive feedback regulation of the KRAS signaling pathway. Currently, the regulation of KRAS-SOS1 interactions and KRAS downstream effector proteins has become a new hotspot in the development of KRAS-driven cancer therapies. However, the detailed dynamic mechanisms of SOS1-catalyzed GDP extraction and the impact of KRAS mutations remain unknown. Herein, the main mechanisms of GDP extraction from KRAS oncogenes by means of the guanine exchange factor SOS1 are disclosed and described with full details at the atomic-level. For GDP-bound wild-type KRAS, four amino acids (Lys811, Glu812, Lys939 and Glu942) responsible for the catalytic function of SOS1 were identified. With the occurrence of KRAS-G12D mutation, the GDP extraction rate is significantly increased. The molecular interactions behind this phenomenon have been subsequently identified being mainly hydrogen bonding interactions between the mutated residue Asp12 and a positively charged pocket located at the intrinsically disordered region807-818 and composed by Ser807, Trp809, Thr810 and Lys811. Our findings provide new insights into the SOS1-KRAS interactions and facilitate the development of related anti-cancer strategies based on the blockage of the above described mechanisms.

biophysics↗

bioRxiv2024

c-di-GMP modulates ribosome assembly by inhibiting rRNA methylation

Cyclic diguanosine monophosphate (c-di-GMP) is a ubiquitous bacterial secondary messenger, with diverse functions, many of which are yet to be uncovered. Stemming from an Escherichia coli proteome microarray, we found that c-di-GMP bound to 23S rRNA methyltransferases (RlmI and RlmE). rRNA methylation assays showed that c-di-GMP inhibits RlmI activity, thereby modulating ribosome assembly. Based on molecular dynamic simulation and mutagenesis studies, we found that c-di-GMP binds to RlmI at residues R64, R103, G114, and K201. Structural simulation revealed that c-di-GMP quenches RlmI activity by inducing the closure of the catalytic pocket. Furthermore, we revealed that c-di-GMP promotes antibiotic tolerance by regulating RlmI activity, which played a role in antibiotic-resistant strains. Finally, the binding and methylation assays showed that the effect of c-di-GMP on RlmI is conserved, at least in various pathogenic bacteria. This study discovered an unexpected functional role of c-di-GMP in regulating ribosome assembly by inhibiting rRNA methylases. This study identified an unexpected but crucial member among the c-di-GMP effectors. HighlightsO_LIc-di-GMP regulates ribosome assembly in Escherichia coli. C_LIO_LIc-di-GMP inhibits rRNA methylation activity of RlmI by inducing catalytic pocket closure. C_LIO_LIc-di-GMP promotes antibiotic resistance by regulating ribosome assembly. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/597503v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@52baforg.highwire.dtl.DTLVardef@176f86dorg.highwire.dtl.DTLVardef@1afed11org.highwire.dtl.DTLVardef@f53687_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2023

Well-tempered metadynamics calculations of free energy surfaces of benzothiadiazine derivatives in aqueous solution

KRAS oncogenes are the largest family of mutated RAS isoforms, participating in about 30% of all cancers. Due to their paramount medical importance, enormous effort is being devoted to the development of inhibitors using clinical tests, wet-lab experiments and drug design, being this a preliminary step in the process of creating new drugs, prior to synthesis and clinical testing. One central aspect in the development of new drugs is the characterization of all species that can be used for treatment. In this aim we propose a computational framework based on combined all-atom molecular dynamics and metadynamics simulations in order to accurately access the most stable conformational variants for several derivatives of a recently proposed small-molecule, called DBD15-21-22. Free energy calculations are essential to unveil mechanisms at the atomic scale like binding affinities or dynamics of stable states. Considering specific atom-atom distances and torsional angles as reliable reaction coordinates we have obtained free-energy landscapes by well-tempered metadynamics simulations, revealing local and global minima of the free-energy hypersurface. We have observed that a variety of stable states together with transitions states are clearly detected depending on the particular species, leading to predictions on the behaviour of such compounds in ionic aqueous solution.

biophysics↗

bioRxiv2023

Molecular Dynamics simulation of NRAS-Q61 oncogenes and new strategies for in silico drug design

The NRAS-mutant subset of melanoma represents the most aggressive and the deadliest types associated with poorer overall survival. Unfortunately, for more than 40 years, no therapeutic agent directly targeting NRAS mutations have been clinically approved yet. Herein, based on microsecond scale molecular dynamics simulations, the concept of NRAS-Q61 mutation classification was firstly proposed. NRAS Q61 positively charged mutations (Q61R and Q61K) was classified together, with a specific targetable pocket, while NRAS-Q61L classified into another category. Moreover, the isomer-sourced structure iteration (ISSI) method was developed for the in silico design of potential inhibitors (HM-516) targeting NRAS mutations. Overall, through this article, we hope to provide the academic and clinical community with new perspective and understanding of NRAS oncoproteins, and propose possible solution to this challenge.

cancer biology↗

bioRxiv2023

Discovery of a new drug using lipid-based formulation targeting G12D mutated KRAS4B through non-covalent bonds

One of the most common drivers in human cancer is KRAS4B. In recent years, the promising KRAS targeted drug development has attracted significant new research interest and reignited the field of RAS therapeutics. To signal, oncogenic KRAS4B not only requires a sufficient nucleotide exchange, but also needs to recruit effectors by exposing its effector-binding sites while anchoring to plasma membrane where KRAS4B-mediated signaling events occur. Phosphodiesterase-{delta} plays an important role in sequestering KRAS4B from the cytoplasm and targeting it to cellular membranes. In this work, we have designed a drug LIG1 using lipid-based formulation to directly target both the switch-II pocket of KRAS4B-G12D and phosphodiesterase-{delta}. LIG1 was found to lock KRAS4B in its GDP-bound state while the effector-binding domain is blocked by the interface of the plasma membrane which hinders the nucleotide exchange while simultaneously it can affect the GTP-bound KRAS4B to shift from an active state to its inactive state. LIG1 is also observed to stably accommodate itself in the prenyl-binding pocket of phosphodiesterase-{delta} which impairs KRAS4B enrichment at the membrane and suppress the proliferation of KRAS4B-dependent cancer cells. In this work we report a drug based on lipid-based formulation that can foster drug discovery efforts for the targeting of oncogenes of the RAS family and beyond.

molecular biology↗

bioRxiv2022

Discovering and targeting dynamic drugging pocketsof the oncogene KRAS-G12D

Activated KRAS-G12D mutations are the one of most frequent oncogenic drivers in human cancers. Unfortunately, no therapeutic agent directly targeting KRAS-G12D has been clinically approved yet, with such mutated species remaining undrugged. Notably, cofactor Mg2+ is closely related to the function of small GTPases, but no investigation has been done yet on Mg2+ when associated with KRAS. Herein, through microsecond scale molecular dynamics simulations we have found that Mg2+ plays a crucial role in the conformational changes of the KRAS-GDP complex. We have located two brand new druggable dynamic pockets exclusive to KRAS-G12D. Using the structural characteristics of these two dynamic pockets, we designed in silico the inhibitor DBD15-21-22, which can specifically and tightly target KRAS-G12D-GDP-Mg2+ ternary complex and we have verified that DBD15-21-22 is harmless for wild-type KRAS. Overall, we provide two brand new druggable pockets located on KRAS-G12D, as well as suitable strategies for KRAS-G12D inhibition.

cancer biology↗

bioRxiv2022

In silico drug design of benzothiadiazine derivatives interactingwith bilayer cell membranes

The use of drugs derived from benzothiadiazine, a bicyclic heterocyclic benzene derivative, has become a widespread treatment for diseases such as hypertension, low blood sugar or the human immunodeficiency virus, among others. In this work we have investigated the interactions of benzothiadiazine and several selected derivatives designed in silico, with the basic components of cell membranes and solvents such as phospholipids, cholesterol and water. The analysis of the mutual microscopic interactions is of central importance to elucidate the local structure of benzothiadiazine as well as the mechanisms responsible for the distribution and access of benzothiadiazine to the interior of the cell. We have performed molecular dynamics simulations of benzothiadiazine and its derivatives embedded in a model zwitterionic bilayer membrane made by phospholipids dioleoylphosphatidylcholine, 1,2-dioleoyl-sn-glycero-3-phosphoserine and cholesterol inside aqueous potassium chloride solution in order to systematically examine microscopic interactions of benzothiadiazine derivatives with the cell membrane at liquid-crystalline phase conditions. From data obtained through radial distribution functions, time dependent hydrogen-bond lengths and potentials of mean force based on reversible work calculations, we have observed that benzothiadiazine derivatives have a strong affinity to stay at the cell membrane interface although their solvation characterisitics can vary significantly: they can be fully solvated by water in short periods of time or continuously attached to specific lipid sites during intervals of 10-70 ns. Furthermore, benzothiadiazines are able to bind lipids and cholesterol chains by means of single and double hydrogen-bonds of different characteristic lengths between 1.6 and 2.1 [A].

biophysics↗

bioRxiv2021

Considerable escape of SARS-CoV-2 variant Omicron to antibody neutralization

The SARS-CoV-2 Omicron variant was first identified in November 2021 in Botswana and South Africa1,2. It has in the meantime spread to many countries and is expected to rapidly become dominant worldwide. The lineage is characterized by the presence of about 32 mutations in the Spike, located mostly in the N-terminal domain (NTD) and the receptor binding domain (RBD), which may enhance viral fitness and allow antibody evasion. Here, we isolated an infectious Omicron virus in Belgium, from a traveller returning from Egypt. We examined its sensitivity to 9 monoclonal antibodies (mAbs) clinically approved or in development3, and to antibodies present in 90 sera from COVID-19 vaccine recipients or convalescent individuals. Omicron was totally or partially resistant to neutralization by all mAbs tested. Sera from Pfizer or AstraZeneca vaccine recipients, sampled 5 months after complete vaccination, barely inhibited Omicron. Sera from COVID-19 convalescent patients collected 6 or 12 months post symptoms displayed low or no neutralizing activity against Omicron. Administration of a booster Pfizer dose as well as vaccination of previously infected individuals generated an anti-Omicron neutralizing response, with titers 5 to 31 fold lower against Omicron than against Delta. Thus, Omicron escapes most therapeutic monoclonal antibodies and to a large extent vaccine-elicited antibodies.

immunology↗

bioRxiv2021

Structure of benzothiadiazine at zwitterionic phospholipid cell membranes

The use of drugs derived from benzothiadiazine, which is a bicyclic heterocyclic benzene derivative, has become a widespread treatment for diseases such as hypertension (treated with diuretics such as bendroflumethiazide or chlorothiazide), low blood sugar (treated with non-diuretic diazoxide) or the human immunodeficiency virus, among others. In this work we have investigated the interactions of benzothiadiazine with the basic components of cell membranes and solvents such as phospholipids, cholesterol, ions and water. The analysis of the mutual microscopic interactions is of central importance to elucidate the local structure of benzothiadiazine as well as the mechanisms responsible for the access of benzothiadiazine to the interior of the cell. We have performed molecular dynamics simulations of benzothiadiazine embedded in three different model zwitterionic bilayer membranes made by dimyristoilphosphatidylcholine, dioleoylphosphatidylcholine, 1,2-dioleoyl-sn-glycero-3-phosphoserine and cholesterol inside aqueous sodium-chloride solution in order to systematically examine microscopic interactions of benzothiadiazine with the cell membrane at liquid-crystalline phase conditions. From data obtained through radial distribution functions, hydrogen-bonding lengths and potentials of mean force based on reversible work calculations, we have observed that benzothiadiazine has a strong affinity to stay at the cell membrane interface although it can be fully solvated by water in short periods of time. Furthermore, benzothiadiazine is able to bind lipids and cholesterol chains by means of single and double hydrogen-bonds of different characteristic lengths.

biophysics↗

bioRxiv2021

Predicting the Conformational Variability of Oncogenic GTP-bound G12D Mutated KRas-4B Proteins at Cell Membranes

KRas proteins are the largest family of mutated Ras isoforms, participating in a wide variety of cancers. Due to their importance, large effort is being carried out on drug development by small-molecule inhibitors. However, understanding protein conformational variability remains a challenge in drug discovery. In the case of the Ras family, their multiple conformational states can affect the binding of potential drug inhibitors. To overcome this challenge, we propose a computational framework based on combined all-atom Molecular Dynamics and Metadynamics simulations able to accurately access conformational variants of the target protein. We tested the methodology using a G12D mutated GTP bound oncogenic KRas-4B protein located at the interface of a DOPC/DOPS/cholesterol model anionic cell membrane. Two main orientations of KRas-4B at the anionic membrane have been obtained and explored. The corresponding angles have been taken as reliable reaction coordinates so that free-energy landscapes have been obtained by well-tempered metadynamics simulations, revealing the local and global minima of KRas-4B binding to the cell membrane, unvealing reactive paths of the system between the two preferential orientations and highlighting opportunities for targeting the unique metastable states through the identification of druggable pockets.

biophysics↗

bioRxiv2020

Long-lasting salt bridges provide the anchoring mechanism of oncogenic KRas-4B at cell membranes

Ras is a family of related proteins participating in all animal cell lineages and organs. Ras proteins work as GDP-GTP binary switches and regulate cytoplasmic signalling networks that are able to control several cellular processes, playing an essential role in signal transduction pathways involved in cell growth, differentiation and survival so that overacting Ras signalling can lead to cancer. One of the hardest challenges to face is, with more than hundred different missense mutations found in cancer, the design of mutation-selective therapeutic strategies. In this work, a G12D mutated farnesylated GTP bound KRas-4B protein has been simulated at the interface of a DOPC/DOPS/cholesterol model anionic cell membrane at the all-atom level. A specific long-lasting salt bridge connection between farnesyl and the hypervariable region of the protein has been identified as the main mechanism responsible of the binding of oncogenic farnesylated KRas-4B to the cell membrane, since this particular bond is absent in both wild-type and oncogenic methylated species of KRas-4B. This finding may lead to a deeper understanding of the mechanisms of protein binding and eventual growing and spreading inside cell membranes. From free energy landscapes obtained by well-tempered metadynamics simulations, we have characterised local and global minima of KRas-4B binding to the cell membrane revealing the main pathways between anchored and released states.

biophysics↗