scholarly journals The importance of the quaternary structure to represent conformational ensembles of the major Mycobacterium tuberculosis drug target

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Renata Fioravanti Tarabini ◽  
Luís Fernando Saraiva Macedo Timmers ◽  
Carlos Eduardo Sequeiros-Borja ◽  
Osmar Norberto de Souza
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Conformational Changes ◽  
Protein Function ◽  
Environmental Changes ◽  
Quaternary Structure ◽  
Single Point ◽  
Point Mutations ◽  
Conformational Ensembles ◽  
Dynamics Simulations ◽  
Coa Reductase

Abstract Flexibility is a feature intimately related to protein function, since conformational changes can be used to describe environmental changes, chemical modifications, protein-protein and protein-ligand interactions. In this study, we have investigated the influence of the quaternary structure of 2-trans-enoyl-ACP (CoA) reductase or InhA, from Mycobacterium tuberculosis, to its flexibility. We carried out classical molecular dynamics simulations using monomeric and tetrameric forms to elucidate the enzyme’s flexibility. Overall, we observed statistically significant differences between conformational ensembles of tertiary and quaternary structures. In addition, the enzyme’s binding site is the most affected region, reinforcing the importance of the quaternary structure to evaluate the binding affinity of small molecules, as well as the effect of single point mutations to InhA protein dynamics.

scholarly journals Modulation of erbB kinase activity and oncogenic potential by single point mutations in the glycine loop of the catalytic domain.

1994 ◽  
Vol 14 (10) ◽  
pp. 6868-6878 ◽  
Author(s):  
H K Shu ◽  
C M Chang ◽  
L Ravi ◽  
L Ling ◽  
C M Castellano ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Conformational Changes ◽  
Kinase Activity ◽  
Catalytic Domain ◽  
Single Point ◽  
Point Mutations ◽  
Full Length ◽  
Site Directed Mutagenesis ◽  
Tyrosine Kinase Domain ◽  
Amino Terminal

Avian c-erbB is activated to a leukemia oncogene following truncation of its amino-terminal ligand-binding domain by retroviral insertion. The insertionally activated transcripts encode protein products which have constitutive tyrosine kinase activity and can induce erythroleukemia but not sarcomas. We have previously found that a valine-to-isoleucine point mutation at position 157 (V157I mutant) within the tyrosine kinase domain of this truncated erbB can dramatically activate the sarcomagenic potential of the oncogene and increase the kinase activity of this oncoprotein. This mutation lies at position 157 of the insertionally activated c-erbB product, affecting a highly conserved valine residue of the glycine loop involved in ATP binding and phosphate transfer. To investigate the functional importance of this residue in the catalytic activity of kinases, we have introduced at this position, by site-directed mutagenesis, codons representing the remaining 18 amino acid residues. Most of the mutants have diminished activity, with six of them completely devoid of kinase activity, indicating the sensitivity of this region to conformational changes. Some of these mutants displayed increased kinase activity and greater transforming potential in comparison with IA c-erbB, but none had levels as high as those of the V157I mutant. In general, the sarcomagenic potential of the various erbB mutants correlated with their autophosphorylation state and their ability to cause phosphorylation of MAP kinase. However, there are important exceptions such as the V157G mutant, which lacks enhanced autophosphorylation but is highly sarcomagenic. Studies of this and other autophosphorylation site mutants point to the existence of an autophosphorylation-independent pathway in sarcomagenesis. The requirement for leukemogenic potential is much less stringent and correlates with positivity of kinase activity. When the valine-to-isoleucine substitution was put in context of the full-length erbB protein, the mutation relaxed the ligand dependence and had a positive effect on the transforming potential of the full-length c-erbB.

scholarly journals Cryo-EM reveals local and global structural rearrangements in RYR mutants

2021 ◽  
Vol 154 (9) ◽  
Author(s):  
Kavita A. Iyer ◽  
Yifan Hu ◽  
Thomas Klose ◽  
Takashi Murayama ◽  
Montserrat Samsó
Keyword(s):  
Long Range ◽  
Conformational Changes ◽  
Large Scale ◽  
Single Point ◽  
Ryanodine Receptors ◽  
Critical Role ◽  
Point Mutations ◽  
Polymorphic Ventricular Tachycardia ◽  
Functional Studies ◽  
Single Point Mutations

Single-point mutations in ryanodine receptors (RYRs), large intracellular Ca2+ channels that play a critical role in EC coupling, are linked to debilitating and lethal disorders such as central core disease, malignant hyperthermia (for the skeletal isoform, RYR1), catecholaminergic polymorphic ventricular tachycardia, and ARVD2 (for the cardiac isoform, RYR2). Mutant RYRs result in elevated [Ca2+]cyto due to steady leak from the sarcoplasmic reticulum. To explore the nature of long-range allosteric mechanisms of malfunction, we determined the structure of two N-terminal domain mutants of RYR1, situated far away from the pore. Cryo-electron microscopy of the N-terminal subdomain A (NTDA) and subdomain C (NTDC) full-length mutants, RYR1 R163C (determined to 3.5 Å resolution), and RYR1 Y522S (determined to 4.0 Å resolution), respectively, reveal large-scale conformational changes in the cytoplasmic assembly under closed-state conditions (i.e., absence of activating Ca2+). The multidomain changes suggest that the mutations induce a preactivated state of the channel in R164C by altering the NTDA+/CD interface, and in Y522S by rearrangement of the α-helical bundle in NTDC. However, the extent of preactivation is considerably higher in Y522S as compared with R163C, which agrees with the increased severity of the Y522S mutation as established by various functional studies. The Y522S mutation represents loss of a spacer residue that is crucial for maintaining optimal orientation of α helices in NTDC, alteration of which has long-range effects felt as far away as ∼100 Å. Additionally, the structure of the Y522S mutant channel under open-state conditions also differs from RYR1 WT open channels. Our developing work with RYR mutants exhibits the diverse mechanisms by which these single-point mutations exert an effect on the channel’s function and highlight the complexity of the multidomain channel, as well as the need for targeted therapies.

scholarly journals Functional determinants of protein assembly into homomeric complexes

2016 ◽  
Author(s):  
L. Therese Bergendahl ◽  
Joseph A. Marsh
Keyword(s):  
Conformational Changes ◽  
Protein Function ◽  
Quaternary Structure ◽  
Systematic Analysis ◽  
Functional Annotations ◽  
Evolutionary Selection ◽  
Large Numbers ◽  
Different Types ◽  
And Function ◽  
The Relationship

AbstractApproximately half of proteins with experimentally determined structures can interact with other copies of themselves and assemble into homomeric complexes, the overwhelming majority of which (>96%) are symmetric. Although homomerisation is often assumed to be functionally beneficial and the result of evolutionary selection, there has been little systematic analysis of the relationship between homomer structure and function. Here, utilizing the large numbers of structures and functional annotations now available, we have investigated how proteins that assemble into different types of homomers are associated with different biological functions. We observe that homomers from different symmetry groups are significantly enriched in distinct functions, and can often provide simple physical and geometrical explanations for these associations in regards to substrate recognition or physical environment. One of the strongest associations is the tendency for metabolic enzymes to form dihedral complexes, which we suggest is closely related to allosteric regulation. We provide a physical explanation for why allostery is related to dihedral complexes: it allows for efficient propagation of conformational changes across isologous (i.e. symmetric) interfaces. Overall we demonstrate a clear relationship between protein function and homomer symmetry that has important implications for understanding protein evolution, as well as for predicting protein function and quaternary structure.

scholarly journals Dynamin-2 R465W mutation induces long range perturbation in highly ordered oligomeric structures

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Fernando Hinostroza ◽  
Alan Neely ◽  
Ingrid Araya-Duran ◽  
Vanessa Marabolí ◽  
Jonathan Canan ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Protein Function ◽  
Molecular Mechanisms ◽  
Protein Complexes ◽  
Coarse Grained ◽  
Cell Physiology ◽  
Missense Mutations ◽  
Small Change ◽  
Dynamics Simulations ◽  
Dynamin 2

Abstract High order oligomers are crucial for normal cell physiology, and protein function perturbed by missense mutations underlies several autosomal dominant diseases. Dynamin-2 is one of such protein forming helical oligomers that catalyze membrane fission. Mutations in this protein, where R465W is the most frequent, cause dominant centronuclear myopathy, but the molecular mechanisms underpinning the functional modifications remain to be investigated. To unveil the structural impact of this mutation in dynamin-2, we used full-atom molecular dynamics simulations and coarse-grained models and built dimers and helices of wild-type (WT) monomers, mutant monomers, or both WT and mutant monomers combined. Our results show that the mutation R465W causes changes in the interactions with neighbor amino acids that propagate through the oligomer. These new interactions perturb the contact between monomers and favor an extended conformation of the bundle signaling element (BSE), a dynamin region that transmits the conformational changes from the GTPase domain to the rest of the protein. This extended configuration of the BSE that is only relevant in the helices illustrates how a small change in the microenvironment surrounding a single residue can propagate through the oligomer structures of dynamin explaining how dominance emerges in large protein complexes.

scholarly journals Mutation-Induced Conformational Changes and Energetics for Binding of FMN Ligand in Flavin Mononucleotide Riboswitch by Molecular Dynamics Simulations

2019 ◽  
Vol 92 (2) ◽  
pp. 241-247
Author(s):  
Padmaja D. Wakchaure ◽  
Bishwajit Ganguly
Keyword(s):  
Molecular Dynamics ◽  
Molecular Dynamics Simulations ◽  
Conformational Changes ◽  
Single Point ◽  
Regulatory Elements ◽  
Flavin Mononucleotide ◽  
Single Point Mutation ◽  
Valuable Insight ◽  
Natural Ligand ◽  
Dynamics Simulations

Riboswitches are the type of regulatory elements present in the untranslated region of mRNA and specifically bind to the natural ligand to regulate gene expression. This binding specificity can be affected by even single point mutation incorporated in the core of the riboswitch. In this work, we have examined the mutations at the binding site residue in Flavin Mononucleotide (FMN) riboswitch structure with 30ns molecular dynamics simulations. The interaction of ligand (FMN) with riboswitch has been characterized using root mean square deviation, hydrogen bonding analysis, and the calculated binding affinities. Mutation at A48G and G62U show the enhanced binding energy however, the mutation at A85G, are energetically unfavorable compared to the wild type. This work gives valuable insight into the structures and energetics of the mutated FMN riboswitch to design new hits for biological applications.

scholarly journals Regulatory Impact of the C-Terminal Tail on Charge Transfer Pathways in Drosophila Cryptochrome

Molecules ◽  
2020 ◽  
Vol 25 (20) ◽  
pp. 4810
Author(s):  
Martin Richter ◽  
Benjamin P. Fingerhut
Keyword(s):  
Charge Transfer ◽  
Conformational Changes ◽  
Quantum Dynamics ◽  
Point Mutations ◽  
Feedback Loops ◽  
Amino Acid Residues ◽  
Linear Interaction ◽  
Regulatory Impact ◽  
Ultrafast Ct ◽  
Dynamics Simulations

Interconnected transcriptional and translational feedback loops are at the core of the molecular mechanism of the circadian clock. Such feedback loops are synchronized to external light entrainment by the blue light photoreceptor cryptochrome (CRY) that undergoes conformational changes upon light absorption by an unknown photoexcitation mechanism. Light-induced charge transfer (CT) reactions in Drosophila CRY (dCRY) are investigated by state-of-the-art simulations that reveal a complex, multi-redox site nature of CT dynamics on the microscopic level. The simulations consider redox-active chromophores of the tryptophan triad (Trp triad) and further account for pathways mediated by W314 and W422 residues proximate to the C-terminal tail (CTT), thus avoiding a pre-bias to specific W-mediated CT pathways. The conducted dissipative quantum dynamics simulations employ microscopically derived model Hamiltonians and display complex and ultrafast CT dynamics on the picosecond timescale, subtly balanced by the electrostatic environment of dCRY. In silicio point mutations provide a microscopic basis for rationalizing particular CT directionality and demonstrate the degree of electrostatic control realized by a discrete set of charged amino acid residues. The predicted participation of CT states in proximity to the CTT relates the directionality of CT reactions to the spatial vicinity of a linear interaction motif. The results stress the importance of CTT directional charge transfer in addition to charge transfer via the Trp triad and call for the use of full-length CRY models including the interactions of photolyase homology region (PHR) and CTT domains.

scholarly journals Coarse-grain simulations on NMR conformational ensembles highlight functional residues in proteins

2019 ◽  
Author(s):  
Sophie Sacquin-Mora
Keyword(s):  
Experimental Data ◽  
Mechanical Properties ◽  
Small Molecules ◽  
Protein Function ◽  
Coarse Grain ◽  
Conformational Ensemble ◽  
Structural Variations ◽  
New Approach ◽  
Conformational Ensembles ◽  
Dynamics Simulations

AbstractDynamics are a key feature of protein function, and this is especially true of gating residues, which occupy cavity or tunnel lining positions in the protein structure, and will reversibly switch between open and closed conformations in order to control the diffusion of small molecules within a protein’s internal matrix. Earlier work on globins and hydrogenases have shown that these gating residues can be detected using a multiscale scheme combining all atom classic molecular dynamics simulations and coarse grain calculations of the resulting conformational ensemble mechanical properties. Here we show that the structural variations observed in the conformational ensembles produced by NMR spectroscopy experiments are sufficient to induce noticeable mechanical changes in a protein, which in turn can be used to identify residues important for function and forming a mechanical nucleus in the protein core. This new approach, which combines experimental data and rapid coarse-grain calculations and no longer needs to resort to time-consuming all-atom simulations, was successfully applied to five different protein families.

scholarly journals Modulation of erbB kinase activity and oncogenic potential by single point mutations in the glycine loop of the catalytic domain

1994 ◽  
Vol 14 (10) ◽  
pp. 6868-6878
Author(s):  
H K Shu ◽  
C M Chang ◽  
L Ravi ◽  
L Ling ◽  
C M Castellano ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Conformational Changes ◽  
Kinase Activity ◽  
Catalytic Domain ◽  
Single Point ◽  
Point Mutations ◽  
Full Length ◽  
Site Directed Mutagenesis ◽  
Tyrosine Kinase Domain ◽  
Amino Terminal

Avian c-erbB is activated to a leukemia oncogene following truncation of its amino-terminal ligand-binding domain by retroviral insertion. The insertionally activated transcripts encode protein products which have constitutive tyrosine kinase activity and can induce erythroleukemia but not sarcomas. We have previously found that a valine-to-isoleucine point mutation at position 157 (V157I mutant) within the tyrosine kinase domain of this truncated erbB can dramatically activate the sarcomagenic potential of the oncogene and increase the kinase activity of this oncoprotein. This mutation lies at position 157 of the insertionally activated c-erbB product, affecting a highly conserved valine residue of the glycine loop involved in ATP binding and phosphate transfer. To investigate the functional importance of this residue in the catalytic activity of kinases, we have introduced at this position, by site-directed mutagenesis, codons representing the remaining 18 amino acid residues. Most of the mutants have diminished activity, with six of them completely devoid of kinase activity, indicating the sensitivity of this region to conformational changes. Some of these mutants displayed increased kinase activity and greater transforming potential in comparison with IA c-erbB, but none had levels as high as those of the V157I mutant. In general, the sarcomagenic potential of the various erbB mutants correlated with their autophosphorylation state and their ability to cause phosphorylation of MAP kinase. However, there are important exceptions such as the V157G mutant, which lacks enhanced autophosphorylation but is highly sarcomagenic. Studies of this and other autophosphorylation site mutants point to the existence of an autophosphorylation-independent pathway in sarcomagenesis. The requirement for leukemogenic potential is much less stringent and correlates with positivity of kinase activity. When the valine-to-isoleucine substitution was put in context of the full-length erbB protein, the mutation relaxed the ligand dependence and had a positive effect on the transforming potential of the full-length c-erbB.

scholarly journals Coarse-grain simulations on NMR conformational ensembles highlight functional residues in proteins

2019 ◽  
Vol 16 (156) ◽  
pp. 20190075
Author(s):  
Sophie Sacquin-Mora
Keyword(s):  
Experimental Data ◽  
Mechanical Properties ◽  
Small Molecules ◽  
Protein Function ◽  
Coarse Grain ◽  
Conformational Ensemble ◽  
Structural Variations ◽  
New Approach ◽  
Conformational Ensembles ◽  
Dynamics Simulations

Dynamics are a key feature of protein function, and this is especially true of gating residues, which occupy cavity or tunnel lining positions in the protein structure, and will reversibly switch between open and closed conformations in order to control the diffusion of small molecules within a protein's internal matrix. Earlier work on globins and hydrogenases have shown that these gating residues can be detected using a multiscale scheme combining all-atom classic molecular dynamics simulations and coarse-grain calculations of the resulting conformational ensemble mechanical properties. Here, we show that the structural variations observed in the conformational ensembles produced by NMR spectroscopy experiments are sufficient to induce noticeable mechanical changes in a protein, which in turn can be used to identify residues important for function and forming a mechanical nucleus in the protein core. This new approach, which combines experimental data and rapid coarse-grain calculations and no longer needs to resort to time-consuming all-atom simulations, was successfully applied to five different protein families.

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