DynaMut: predicting the impact of mutations on protein conformation, flexibility and stability

Rodrigues, Carlos H M; Pires, Douglas E V; Ascher, David B.

doi:10.1093/nar/gky300

Cited by 835 publications

(734 citation statements)

References 47 publications

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“…The different residues located near the inter‐domain region could be responsible for the slightly different conformations of both chitinases, the differences in the kinetic properties and also the different thermodynamic parameters, which could promote divergence in the formation of the protein–ligand complex. In this regard, it has been recognized that mutations can exert long‐distance effects on protein structure, flexibility and stability . Finally, we also demonstrated that both enzymes, one of which (AtChi2) was obtained from RNA of F. oxysporum‐ infected plants, exhibit antifungal activity versus Aspergillus sp.…”

Section: Discussionmentioning

confidence: 53%

A biophysical and structural study of two chitinases from Agave tequilana and their potential role as defense proteins

et al. 2019

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Plant chitinases are enzymes that have several functions, including providing protection against pathogens. Agave tequilana is an economically important plant that is poorly studied. Here, we identified a chitinase from short reads of the A. tequilana transcriptome (AtChi1). A second chitinase, differing by only six residues from the first, was isolated from total RNA of plants infected with Fusarium oxysporum (AtChi2). Both enzymes were overexpressed in Escherichia coli and analysis of their sequences indicated that they belong to the class I glycoside hydrolase family19, whose members exhibit two domains: a carbohydrate‐binding module and a catalytic domain, connected by a flexible linker. Activity assays and thermal shift experiments demonstrated that the recombinant Agave enzymes are highly thermostable acidic endochitinases with Tm values of 75 °C and 71 °C. Both exhibit a molecular mass close to 32 kDa, as determined by MALDI‐TOF, and experimental pIs of 3.7 and 3.9. Coupling small‐angle x‐ray scattering information with homology modeling and docking simulations allowed us to structurally characterize both chitinases, which notably show different interactions in the binding groove. Even when the six different amino acids are all exposed to solvent in the loops located near the linker and opposite to the binding site, they confer distinct kinetic parameters against colloidal chitin and similar affinity for (GlnNAc)6, as shown by isothermal titration calorimetry. Interestingly, binding is more enthalpy‐driven for AtChi2. Whereas the physiological role of these chitinases remains unknown, we demonstrate that they exhibit important antifungal activity against chitin‐rich fungi such as Aspergillus sp. Database SAXS structural data are available in the SASBDB database with accession numbers SASDDE7 and SASDDA6. Enzymes Chitinases (http://www.chem.qmul.ac.uk/iubmb/enzyme/EC3/2/1/14.html).

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Section: Discussionmentioning

confidence: 53%

A biophysical and structural study of two chitinases from Agave tequilana and their potential role as defense proteins

et al. 2019

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“…The coordinates were submitted to the DynaMut server for analysis. 24 For molecular dynamics (MD), allatom models of human TMEM175 were constructed with MODELLER, 25 using the crystal structure of Chamaesiphon minutus TMEM175 channel 26 (protein data bank ID: pdb 5VRE) as a template. For chain A and chain B, residues 36 to 230 and 261 to 473 were aligned using LOMETS 27 ; this returns 10 pairwise alignments, which we combined into a single alignment for which no gaps were introduced in alpha-helices (see Fig 1A).…”

Section: Homology Modeling and Molecular Dynamics Simulationsmentioning

confidence: 99%

Genetic, Structural, and Functional Evidence Link TMEM175 to Synucleinopathies

Krohn

Öztürk

Vanderperre

et al. 2019

Annals of Neurology

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Objective The TMEM175/GAK/DGKQ locus is the 3rd strongest risk locus in genome‐wide association studies of Parkinson disease (PD). We aimed to identify the specific disease‐associated variants in this locus, and their potential implications. Methods Full sequencing of TMEM175/GAK/DGKQ followed by genotyping of specific associated variants was performed in PD (n = 1,575) and rapid eye movement sleep behavior disorder (RBD) patients (n = 533) and in controls (n = 1,583). Adjusted regression models and a meta‐analysis were performed. Association between variants and glucocerebrosidase (GCase) activity was analyzed in 715 individuals with available data. Homology modeling, molecular dynamics simulations, and lysosomal localization experiments were performed on TMEM175 variants to determine their potential effects on structure and function. Results Two coding variants, TMEM175 p.M393T (odds ratio [OR] = 1.37, p = 0.0003) and p.Q65P (OR = 0.72, p = 0.005), were associated with PD, and p.M393T was also associated with RBD (OR = 1.59, p = 0.001). TMEM175 p.M393T was associated with reduced GCase activity. Homology modeling and normal mode analysis demonstrated that TMEM175 p.M393T creates a polar side‐chain in the hydrophobic core of the transmembrane, which could destabilize the domain and thus impair either its assembly, maturation, or trafficking. Molecular dynamics simulations demonstrated that the p.Q65P variant may increase stability and ion conductance of the transmembrane protein, and lysosomal localization was not affected by these variants. Interpretation Coding variants in TMEM175 are likely to be responsible for the association in the TMEM175/GAK/DGKQ locus, which could be mediated by affecting GCase activity. ANN NEUROL 2020;87:139–153

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“…The arginine-323 to methionine mutation was modeled by selecting the most probable rotamer conformation according to the Richardson backbone-independent rotamer library (Lovell et al 2000). The predicted effects of the R323C mutation on TRMT1 structure was performed using DynaMut (Rodrigues et al 2018).…”

Section: In Silico Analysis Of Trmt1 Structurementioning

confidence: 99%

A missense variant impairing TRMT1 function in tRNA modification is linked to intellectual disability

Zhang

Lentini

Prevost

et al. 2019

Preprint

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The human TRMT1 gene encodes a tRNA methyltransferase enzyme responsible for the formation of the dimethylguanosine (m2,2G) modification in cytoplasmic and mitochondrial tRNAs. Frameshift mutations in the TRMT1 gene have been shown to cause autosomal-recessive intellectual disability (ID) in the human population but additional TRMT1 variants remain to be characterized. Moreover, the impact of ID-associated TRMT1 mutations on m2,2G levels in IDaffected patients is unknown. Here, we describe a homozygous missense variant in TRMT1 in a patient displaying developmental delay, ID, and epilepsy. The missense variant changes a conserved arginine residue to a cysteine (R323C) within the methyltransferase domain of TRMT1 and is expected to perturb protein folding. Patient cells expressing the TRMT1-R323C variant exhibit a severe deficiency in m2,2G modifications within tRNAs, indicating that the mutation causes loss-of-function. Notably, the TRMT1 R323C mutant retains the ability to bind tRNA but is unable to rescue m2,2G formation in TRMT1-deficient human cells. Our results identify a pathogenic point mutation in TRMT1 that severely perturbs tRNA modification activity, and provide the first demonstration that m2,2G modifications are disrupted in patients with TRMT1associated ID disorders.

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DynaMut: predicting the impact of mutations on protein conformation, flexibility and stability

Cited by 835 publications

References 47 publications

A biophysical and structural study of two chitinases from Agave tequilana and their potential role as defense proteins

A biophysical and structural study of two chitinases from Agave tequilana and their potential role as defense proteins

Genetic, Structural, and Functional Evidence Link TMEM175 to Synucleinopathies

A missense variant impairing TRMT1 function in tRNA modification is linked to intellectual disability

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