Reversibility and two state behaviour in the thermal unfolding of oligomeric TIM barrel proteins

Romero‐Romero, Sergio; Costas, Miguel; Rodrı́guez-Romero, Adela; Fernández‐Velasco, D. Alejandro

doi:10.1039/c5cp01599e

Cited by 22 publications

(35 citation statements)

References 123 publications

(303 reference statements)

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“…For Sc TIM the discontinuous line represents a simulation with a K diss of 1 × 10 −11 m . Discontinuous line for Bac TIM represents a simulation for the bacterial TIM from Streptomyces coelicolor with a K diss of 1.8 × 10 −7 m . (C) Thermal unfolding of anc TIM s followed by CD.…”

Section: Resultsmentioning

confidence: 99%

“…The coupling between association, stability and catalysis in TIM has been previously addressed by studying extant enzymes from a variety of sources, using site directed mutagenesis, computational design, directed evolution and consensus design . In this work, we addressed the evolution of this coupling by ASR.…”

Section: Introductionmentioning

confidence: 99%

“…In this work, we addressed the evolution of this coupling by ASR. Only some extant bacterial TIMs have been characterized in detail and archaeal TIMs are shorter than those from the other domains and show changes in the oligomerization state . In order to increase the confidence of the reconstruction process and to compare the results obtained with those previously reported for extant TIMs, we decided to focus on eukaryotic TIMs.…”

Section: Introductionmentioning

confidence: 99%

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Structural, thermodynamic and catalytic characterization of an ancestral triosephosphate isomerase reveal early evolutionary coupling between monomer association and function

et al. 2019

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Function, structure, and stability are strongly coupled in obligated oligomers, such as triosephosphate isomerase (TIM). However, little is known about how this coupling evolved. To address this question, five ancestral TIMs (ancTIMs) in the opisthokont lineage were inferred. The encoded proteins were purified and characterized, and spectroscopic and hydrodynamic analysis indicated that all are folded dimers. The catalytic efficiency of ancTIMs is very high and all dissociate into inactive and partially unfolded monomers. The placement of catalytic residues in the three‐dimensional structure, as well as the enthalpy‐driven binding signature of the oldest ancestor (TIM63) resemble extant TIMs. Although TIM63 dimers dissociate more readily than do extant TIMs, calorimetric data show that the free ancestral subunits are folded to a greater extent than their extant counterparts are, suggesting that full catalytic proficiency was established in the dimer before the stability of the isolated monomer eroded. Notably, the low association energy in ancTIMs is compensated for by a high activation barrier, and by a significant shift in the dimer‐monomer equilibrium induced by ligand binding. Our results indicate that before the animal and fungi lineages diverged, TIM was an obligated oligomer with substrate binding properties and catalytic efficiency that resemble that of extant TIMs. Therefore, TIM function and association have been strongly coupled at least for the last third of biological evolution on earth. Databases PDB Entry: http://www.rcsb.org/pdb/search/structidSearch.do?structureId=6NEE. Enzymes Triosephosphate isomerase http://www.chem.qmul.ac.uk/iubmb/enzyme/EC5/3/1/1.html, Glycerol‐3‐phosphate dehydrogenase http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/1/1/8.html.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Structural, thermodynamic and catalytic characterization of an ancestral triosephosphate isomerase reveal early evolutionary coupling between monomer association and function

et al. 2019

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“…As TPI is a homodimer, a decrease in total protein concentration favors dimer dissociation. app K diss was calculated from the following equation, derived from the simplest equilibrium two‐state model that describes the change in the fraction of active dimers f N as a function of TPI concentration ( P C expressed on a monomer basis) (Neet and Timm, ; Romero‐Romero et al ., )

f_{N} = \frac{^{app} K_{diss} + 4 P_{C} - \sqrt{^{app} {K_{diss}}^{2} + 8 K_{normaldiss} P_{C}}}{4 P_{C}}

…”

Section: Methodsmentioning

confidence: 99%

Structural basis for the modulation of plant cytosolic triosephosphate isomerase activity by mimicry of redox‐based modifications

et al. 2019

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Summary Reactive oxidative species (ROS) and S‐glutathionylation modulate the activity of plant cytosolic triosephosphate isomerases (cTPI). Arabidopsis thaliana cTPI (AtcTPI) is subject of redox regulation at two reactive cysteines that function as thiol switches. Here we investigate the role of these residues, AtcTPI‐Cys13 and At‐Cys218, by substituting them with aspartic acid that mimics the irreversible oxidation of cysteine to sulfinic acid and with amino acids that mimic thiol conjugation. Crystallographic studies show that mimicking AtcTPI‐Cys13 oxidation promotes the formation of inactive monomers by reposition residue Phe75 of the neighboring subunit, into a conformation that destabilizes the dimer interface. Mutations in residue AtcTPI‐Cys218 to Asp, Lys, or Tyr generate TPI variants with a decreased enzymatic activity by creating structural modifications in two loops (loop 7 and loop 6) whose integrity is necessary to assemble the active site. In contrast with mutations in residue AtcTPI‐Cys13, mutations in AtcTPI‐Cys218 do not alter the dimeric nature of AtcTPI. Therefore, modifications of residues AtcTPI‐Cys13 and AtcTPI‐Cys218 modulate AtcTPI activity by inducing the formation of inactive monomers and by altering the active site of the dimeric enzyme, respectively. The identity of residue AtcTPI‐Cys218 is conserved in the majority of plant cytosolic TPIs, this conservation and its solvent‐exposed localization make it the most probable target for TPI regulation upon oxidative damage by reactive oxygen species. Our data reveal the structural mechanisms by which S‐glutathionylation protects AtcTPI from irreversible chemical modifications and re‐routes carbon metabolism to the pentose phosphate pathway to decrease oxidative stress.

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“…Being able to relate the pathogenic characteristics of each of these organisms and the enzymatic activity of its TIM, it is what can help us to find a consensus region. The most susceptible organisms such as T. vaginalis, C. perfringens and S. coelicolor, have specific characteristics, that can give the difference to its TIM, where, extremophilic bacteria such as T. thermophiles and D. radiodurans , with characteristics that make them unique to resist extreme conditions, temperatures, ionizing radiation, reactive oxygen species and repair denatured proteins . These differences in the organisms and residues sequences of the TIMs give them specific regions that will help us to choose a potential interaction site for compounds.…”

Section: Introductionmentioning

confidence: 99%

Potential Site to Direct Selective Compounds in the Triosephosphate Isomerase for the Development of New Drugs

et al. 2020

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In the pharmaceutical industry, the development of selective drugs to an enzyme or the repositioning of commercial drugs, today, is booming. The glycolytic enzyme triosephosphate isomerase (TIM) has been used as a therapeutic target for the development of new drugs against various pathogenic organ-isms. Therefore, saving resources in the development of new drugs, by directing the interaction of pharmacological compounds to an interaction site with a high probability to be selective, represents an opportunity for researchers.In this study, we propose a potential site as a therapeutic target against TIM, for the development of drugs with probability to be safe in humans.We propose that K214, which is in the position near the sequence Y209, G210, G211, S212, V213, is indispensable for interaction with the tested compounds to interact near the active site of TIMs. In addition, the TIMs that present F46, achieve a better interaction and a greater effect on the decrease in the enzymatic activity of the compounds tested. Therefore, we determine compounds with this effect and from its derivatives could be used in other TIMs. To contribute to the development of new selective drugs against bacteria, parasites or other organisms that nowadays have non-specific conventional treatments.

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Reversibility and two state behaviour in the thermal unfolding of oligomeric TIM barrel proteins

Cited by 22 publications

References 123 publications

Structural, thermodynamic and catalytic characterization of an ancestral triosephosphate isomerase reveal early evolutionary coupling between monomer association and function

Structural, thermodynamic and catalytic characterization of an ancestral triosephosphate isomerase reveal early evolutionary coupling between monomer association and function

Structural basis for the modulation of plant cytosolic triosephosphate isomerase activity by mimicry of redox‐based modifications

Potential Site to Direct Selective Compounds in the Triosephosphate Isomerase for the Development of New Drugs

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