Ensemble Perspective for Catalytic Promiscuity

Honaker, Matthew T.; Acchione, Mauro; Sumida, John P.; Atkins, William M.

doi:10.1074/jbc.m111.304386

Cited by 25 publications

(9 citation statements)

References 53 publications

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“…The functional dimer found in GSTs was generated by the symmetry operator -x, y, -z+½ of the C 222 1 space group (Figure 3B). The interface involves 49 residues from each monomer and the buried surface area is ∼1645 Å 2 from each monomer (about 15% of the total solvent accessible area of each monomer), which is within the values found in most other GST families [1], [7], [9], [10]. The main regions involved in subunit interactions are residues 65–72 (part of helix H3), 80–85 (strand β4), 86–96 (helix H4), 105–128 (part of helix H5), 139–143 and 154–162 from helix H6.…”

Section: Resultssupporting

confidence: 74%

“…GSTs are known as promiscuous enzymes capable of catalyzing the conjugation of GSH with a broad range of electrophilic substrates [5]–[7]. Several members of the GST family are selectively induced by biotic and abiotic stress treatments and play important roles in the regulation of redox homeostasis as well as in endogenous metabolism [3], [4].…”

Section: Introductionmentioning

confidence: 99%

“…Glutathione transferases (GSTs; EC 2.5.1.18) are phase II detoxification enzymes that metabolize a wide range of hydrophobic toxic compounds by catalyzing the conjugation of glutathione (GSH) to the hydrophilic centre of the toxic substances [1] – [4] . GSTs are known as promiscuous enzymes capable of catalyzing the conjugation of GSH with a broad range of electrophilic substrates [5] – [7] . Several members of the GST family are selectively induced by biotic and abiotic stress treatments and play important roles in the regulation of redox homeostasis as well as in endogenous metabolism [3] , [4] .…”

Section: Introductionmentioning

confidence: 99%

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A Glutathione Transferase from Agrobacterium tumefaciens Reveals a Novel Class of Bacterial GST Superfamily

et al. 2012

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In the present work, we report a novel class of glutathione transferases (GSTs) originated from the pathogenic soil bacterium Agrobacterium tumefaciens C58, with structural and catalytic properties not observed previously in prokaryotic and eukaryotic GST isoenzymes. A GST-like sequence from A. tumefaciens C58 (Atu3701) with low similarity to other characterized GST family of enzymes was identified. Phylogenetic analysis showed that it belongs to a distinct GST class not previously described and restricted only in soil bacteria, called the Eta class (H). This enzyme (designated as AtuGSTH1-1) was cloned and expressed in E. coli and its structural and catalytic properties were investigated. Functional analysis showed that AtuGSTH1-1 exhibits significant transferase activity against the common substrates aryl halides, as well as very high peroxidase activity towards organic hydroperoxides. The crystal structure of AtuGSTH1-1 was determined at 1.4 Å resolution in complex with S-(p-nitrobenzyl)-glutathione (Nb-GSH). Although AtuGSTH1-1 adopts the canonical GST fold, sequence and structural characteristics distinct from previously characterized GSTs were identified. The absence of the classic catalytic essential residues (Tyr, Ser, Cys) distinguishes AtuGSTH1-1 from all other cytosolic GSTs of known structure and function. Site-directed mutagenesis showed that instead of the classic catalytic residues, an Arg residue (Arg34), an electron-sharing network, and a bridge of a network of water molecules may form the basis of the catalytic mechanism. Comparative sequence analysis, structural information, and site-directed mutagenesis in combination with kinetic analysis showed that Phe22, Ser25, and Arg187 are additional important residues for the enzyme's catalytic efficiency and specificity.

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Section: Resultssupporting

confidence: 74%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A Glutathione Transferase from Agrobacterium tumefaciens Reveals a Novel Class of Bacterial GST Superfamily

et al. 2012

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“…This hypothesis is difficult to address without tools to quantify promiscuity and ‘flexibility’. Honaker et al [65,66] used a series of mutants of the highly promiscuous GST enzymes to determine their substrate promiscuity against a highly diverse basis set of substrates. Fortuitously, some A-class GSTs exhibited a reversible low temperature, barrierless transition in differential scanning calorimetry (DSC) experiments that corresponds to rearrangement of the active site and its C-terminal ‘lid.’ From a specialized analysis of the DSC transition it was possible to estimate the conformational breadth or diversity of the active sites for the series of GSTs with a thermodynamic treatment.…”

Section: Methods For Measuring Promiscuitymentioning

confidence: 99%

Biological messiness vs. biological genius: Mechanistic aspects and roles of protein promiscuity

Atkins

2015

The Journal of Steroid Biochemistry and Molecular Biology

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In contrast to the traditional biological paradigms focused on ‘specificity’, recent research and theoretical efforts have focused on functional ‘promiscuity’ exhibited by proteins and enzymes in many biological settings, including enzymatic detoxication, steroid biochemistry, signal transduction and immune responses. In addition, divergent evolutionary processes are apparently facilitated by random mutations that yield promiscuous enzyme intermediates. The intermediates, in turn, provide opportunities for further evolution to optimize new functions from existing protein scaffolds. In some cases, promiscuity may simply represent the inherent plasticity of proteins resulting from their polymeric nature with distributed conformational ensembles. Enzymes or proteins that bind or metabolize noncognate substrates create ‘messiness’ or noise in the systems they contribute to. With our increasing awareness of the frequency of these promiscuous behaviors it becomes interesting and important to understand the molecular bases for promiscuous behavior and to distinguish between evolutionarily selected promiscuity and evolutionarily tolerated messiness. This review provides an overview of current understanding of these aspects of protein biochemistry and enzymology.

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“…Another example is the antiviral protein RhTC, which was found to target different HIV viruses by allowing a dynamic active site to assume very different conformations [16]. More generally, emerging evidence is lending support to the view that functional promiscuity in enzymes may frequently be based on thermodynamic fluctuations of conformational sub-states [17]. However, this may not always be the case, for example, if the functional promiscuity is mediated by changes of catalytic residues that do not cause conformational changes.…”

Section: Introductionmentioning

confidence: 99%

Evolutionary Dynamics on Protein Bi-stability Landscapes can Potentially Resolve Adaptive Conflicts

2012

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Experimental studies have shown that some proteins exist in two alternative native-state conformations. It has been proposed that such bi-stable proteins can potentially function as evolutionary bridges at the interface between two neutral networks of protein sequences that fold uniquely into the two different native conformations. Under adaptive conflict scenarios, bi-stable proteins may be of particular advantage if they simultaneously provide two beneficial biological functions. However, computational models that simulate protein structure evolution do not yet recognize the importance of bi-stability. Here we use a biophysical model to analyze sequence space to identify bi-stable or multi-stable proteins with two or more equally stable native-state structures. The inclusion of such proteins enhances phenotype connectivity between neutral networks in sequence space. Consideration of the sequence space neighborhood of bridge proteins revealed that bi-stability decreases gradually with each mutation that takes the sequence further away from an exactly bi-stable protein. With relaxed selection pressures, we found that bi-stable proteins in our model are highly successful under simulated adaptive conflict. Inspired by these model predictions, we developed a method to identify real proteins in the PDB with bridge-like properties, and have verified a clear bi-stability gradient for a series of mutants studied by Alexander et al. (Proc Nat Acad Sci USA 2009, 106:21149–21154) that connect two sequences that fold uniquely into two different native structures via a bridge-like intermediate mutant sequence. Based on these findings, new testable predictions for future studies on protein bi-stability and evolution are discussed.

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Ensemble Perspective for Catalytic Promiscuity

Cited by 25 publications

References 53 publications

A Glutathione Transferase from Agrobacterium tumefaciens Reveals a Novel Class of Bacterial GST Superfamily

A Glutathione Transferase from Agrobacterium tumefaciens Reveals a Novel Class of Bacterial GST Superfamily

Biological messiness vs. biological genius: Mechanistic aspects and roles of protein promiscuity

Evolutionary Dynamics on Protein Bi-stability Landscapes can Potentially Resolve Adaptive Conflicts

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