Phosphopeptide binding to the N-SH2 domain of tyrosine phosphatase SHP2 correlates with the unzipping of its central β-sheet.

SHP2 is a tyrosine phosphatase that plays a regulatory role in multiple intracellular signaling cascades and is known to be oncogenic in certain contexts. In the absence of effectors, SHP2 adopts an autoinhibited conformation with its N-SH2 domain blocking the active site. Given the key role of N-SH2 in regulating SHP2, this domain has been extensively studied, often by X-ray crystallography.

Atomistic ensemble of active SHP2 phosphatase.

SHP2 phosphatase plays an important role in regulating several intracellular signaling pathways. Pathogenic mutations of SHP2 cause developmental disorders and are linked to hematological malignancies and cancer. SHP2 comprises two tandemly-arranged SH2 domains, a catalytic PTP domain, and a disordered C-terminal tail.

Revealing Allostery in PTPN11 SH2 Domains from MD Simulations.

Src-homology 2 (SH2) domains are protein interaction domains that bind to specific peptide motifs containing phosphotyrosine. SHP2, a tyrosine phosphatase encoded by PTPN11 gene, which has been emerged as positive or negative modulator in multiple signaling pathways, contains two SH2 domains, respectively, called N-SH2 and C-SH2. These domains play a relevant role in regulating SHP2 activity, either by recognizing its binding partners or by blocking its catalytic site.

Probing the Role of Murine Neuroglobin CDloop-D-Helix Unit in CO Ligand Binding and Structural Dynamics.

We produced a neuroglobin variant, namely, Ngb CDless, with the excised CDloop- and D-helix, directly joining the C- and E-helices. The CDless variant retained bis-His hexacoordination, and we investigated the role of the CDloop-D-helix unit in controlling the CO binding and structural dynamics by an integrative approach based on X-ray crystallography, rapid mixing, laser flash photolysis, resonance Raman spectroscopy, and molecular dynamics simulations. Rapid mixing and laser flash photolysis showed that ligand affinity was unchanged with respect to the wild-type protein, albeit with increased on and off constants for rate-limiting heme iron hexacoordination by the distal His64.

Conformational dynamics promotes disordered regions from function-dispensable to essential in evolved site-specific DNA recombinases.

Protein intrinsically disordered regions (IDRs) play pivotal roles in molecular recognition and regulatory processes through structural disorder-to-order transitions. To understand and exploit the distinctive functional implications of IDRs and to unravel the underlying molecular mechanisms, structural disorder-to-function relationships need to be deciphered. The DNA site-specific recombinase system Cre/loxP represents an attractive model to investigate functional molecular mechanisms of IDRs.

The loops of the N-SH2 binding cleft do not serve as allosteric switch in SHP2 activation.

The Src-homology-2 domain-containing phosphatase SHP2 is a critical regulator of signal transduction, being implicated in cell growth and differentiation. Activating mutations cause developmental disorders and act as oncogenic drivers in hematologic cancers. SHP2 is activated by phosphopeptide binding to the N-SH2 domain, triggering the release of N-SH2 from the catalytic PTP domain.

An allosteric interaction controls the activation mechanism of SHP2 tyrosine phosphatase.

SHP2 is a protein tyrosine phosphatase (PTP) involved in multiple signaling pathways. Mutations of SHP2 can result in Noonan syndrome or pediatric malignancies. Inhibition of wild-type SHP2 represents a novel strategy against several cancers.

Pathogenic PTPN11 variants involving the poly-glutamine Gln -Gln -Gln stretch highlight the relevance of helix B in SHP2's functional regulation.

Germline PTPN11 mutations cause Noonan syndrome (NS), the most common disorder among RASopathies. PTPN11 encodes SHP2, a protein tyrosine-phosphatase controlling signaling through the RAS-MAPK and PI3K-AKT pathways. Generally, NS-causing PTPN11 mutations are missense changes destabilizing the inactive conformation of the protein or enhancing its binding to signaling partners.

Proximal and distal control for ligand binding in neuroglobin: role of the CD loop and evidence for His64 gating.

Neuroglobin (Ngb) is predominantly expressed in neurons of the central and peripheral nervous systems and it clearly seems to be involved in neuroprotection. Engineering Ngb to observe structural and dynamic alterations associated with perturbation in ligand binding might reveal important structural determinants, and could shed light on key features related to its mechanism of action. Our results highlight the relevance of the CD loop and of Phe106 as distal and proximal controls involved in ligand binding in murine neuroglobin.

Dynamic multiple-scattering treatment of X-ray absorption: Parameterization of a new molecular dynamics force field for myoglobin.

We present a detailed analysis of the X-ray absorption near-edge structure (XANES) data on the Fe K-edge of CO Myoglobin based on a combined procedure of Molecular Dynamics (MD) calculations and MXAN (Minuit XANes) data analysis that we call D-MXAN. The ability of performing quantitative XANES data analysis allows us to refine classical force field MD parameters, thus obtaining a reliable tool for the atomic investigation of this important model system for biological macromolecules. The iterative procedure here applied corrects the greatest part of the structural discrepancy between classical MD sampling and experimental determinations.

Clinical and functional characterization of two novel ZBTB20 mutations causing Primrose syndrome.

Primrose syndrome (PS) is a rare disorder characterized by macrocephaly, tall stature, intellectual disability, autistic traits, and disturbances of glucose metabolism with insulin-resistant diabetes and distal muscle wasting occurring in adulthood. The disorder is caused by functional dysregulation of ZBTB20, a transcriptional repressor controlling energetic metabolism and developmental programs. ZBTB20 maps in a genomic region that is deleted in the 3q13.

Exploring Multiple Binding Modes Using Confined Replica Exchange Molecular Dynamics.

Molecular docking is extensively applied to determine the position of a ligand on its receptor despite the rather poor correspondence between docking scores and experimental binding affinities found in several studies, especially for systems structurally unrelated with those used in the scoring functions' training sets. Here, we present a method for the prediction of binding modes and binding free energies, which uses replica exchange molecular dynamics in combination with a receptor-shaped piecewise potential, confining the ligand in the proximity of the receptor surface and limiting the accessible conformational space of interest. We assess our methodology with a set of protein receptor-ligand test cases.

Mutations Impairing GSK3-Mediated MAF Phosphorylation Cause Cataract, Deafness, Intellectual Disability, Seizures, and a Down Syndrome-like Facies.

Transcription factors operate in developmental processes to mediate inductive events and cell competence, and perturbation of their function or regulation can dramatically affect morphogenesis, organogenesis, and growth. We report that a narrow spectrum of amino-acid substitutions within the transactivation domain of the v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog (MAF), a leucine zipper-containing transcription factor of the AP1 superfamily, profoundly affect development. Seven different de novo missense mutations involving conserved residues of the four GSK3 phosphorylation motifs were identified in eight unrelated individuals.

Sulfated glycosaminoglycans exploit the conformational plasticity of bone morphogenetic protein-2 (BMP-2) and alter the interaction profile with its receptor.

Sulfated glycosaminoglycans (GAGs) can direct cellular processes by interacting with proteins of the extracellular matrix (ECM). In this study we characterize the interaction profiles of chemically sulfated hyaluronan (HA) and chondroitin sulfate (CS) derivatives with bone morphogenetic protein-2 (BMP-2) and investigate their relevance for complex formation with the receptor BMPR-IA. These goals were addressed by surface plasmon resonance (SPR) and ELISA in combination with molecular modeling and dynamics simulation.

Mutations in ZBTB20 cause Primrose syndrome.

Primrose syndrome and 3q13.31 microdeletion syndrome are clinically related disorders characterized by tall stature, macrocephaly, intellectual disability, disturbed behavior and unusual facial features, with diabetes, deafness, progressive muscle wasting and ectopic calcifications specifically occurring in the former. We report that missense mutations in ZBTB20, residing within the 3q13.

Analysis of infrared spectra of β-hairpin peptides as derived from molecular dynamics simulations.

Infrared temperature-dependent spectroscopy is a well-known tool to characterize folding/unfolding transitions in peptides and proteins, assuming that the higher the temperature, the higher the unfolded population. The infrared spectra at different temperatures of two β-hairpin peptides (gramicidin S analogues GS6 and GS10) are here reconstructed by means of molecular dynamics (MD) simulations and a theoretical-computational method based on the perturbed matrix method. The calculated temperature-dependent spectra result in good agreement with the experimental available spectra.

Structure of the lipodepsipeptide syringomycin E in phospholipids and sodium dodecylsulphate micelle studied by circular dichroism, NMR spectroscopy and molecular dynamics.

Syringomycin E (SRE) is a member of a family of lipodepsipeptides that characterize the secondary metabolism of the plant-associated bacteria Pseudomonas syringae pv. syringae. It displays phytotoxic, antifungal and haemolytic activities, due to the membrane interaction and ion channel formation.

Kinetics of carbon monoxide migration and binding in solvated neuroglobin as revealed by molecular dynamics simulations and quantum mechanical calculations.

Neuroglobin (Ngb) is a globular protein that reversibly binds small ligands at the six coordination position of the heme. With respect to other globins similar to myoglobin, Ngb displays some peculiarities as the topological reorganization of the internal cavities coupled to the sliding of the heme, or the binding of the endogenous distal histidine to the heme in the absence of an exogenous ligand. In this Article, by using multiple (independent) molecular dynamics trajectories (about 500 ns in total), the migration pathways of photolized carbon monoxide (CO) within solvated Ngb were analyzed, and a quantitative description of CO migration and corresponding kinetics was obtained.

The effects of the L29F mutation on the ligand migration kinetics in crystallized myoglobin as revealed by molecular dynamics simulations.

By using multiple molecular dynamics (MD) trajectories, a quantitative description of carbon monoxide (CO) migration within crystal of L29F myoglobin mutant (L29F-Mb) was obtained. The aim was to provide a detailed model for ligand diffusion in the protein to be compared to the available L29F-Mb experimental-computational data and to the corresponding model kinetics we previously obtained for photolyzed CO within crystallized wild-type myoglobin (wt-Mb). Results suggest a clear migration pathway from distal pocket to the proximal site, similar to the one observed in wt-Mb, with a relaxation kinetics differing from the wt-Mb one essentially for the escape rate which is much higher in the mutant.

Dynamic investigation of protein metal active sites: interplay of XANES and molecular dynamics simulations.

The effect of structural disorder on the X-ray absorption near-edge structure (XANES) spectrum of a heme protein has been investigated using the dynamical description of the system derived from molecular dynamics (MD) simulations. The XANES spectra of neuroglobin (Ngb) and carbonmonoxy-neuroglobin (NgbCO) have been quantitatively reproduced, starting from the MD geometrical configurations, without carrying out any optimization in the structural parameter space. These results provide an important experimental validation of the reliability of the potentials used in the MD simulations and accordingly corroborate the consistency of the structural dynamic information on the metal center, related to its biological function.

Combining crystallography and molecular dynamics: the case of Schistosoma mansoni phospholipid glutathione peroxidase.

Oxidative stress is a widespread challenge for living organisms, and especially so for parasitic ones, given the fact that their hosts can produce reactive oxygen species (ROS) as a mechanism of defense. Thus, long lived parasites, such as the flatworm Schistosomes, have evolved refined enzymatic systems capable of detoxifying ROS. Among these, glutathione peroxidases (Gpx) are a family of sulfur or selenium-dependent isozymes sharing the ability to reduce peroxides using the reducing equivalents provided by glutathione or possibly small proteins such as thioredoxin.