The Hydrophobic Region of the DmsA Twin-Arginine Leader Peptide Determines Specificity with Chaperone DmsD

Winstone, Tara M.L.; Tran, Vy A.; Turner, Raymond J.

doi:10.1021/bi4009374

Cited by 16 publications

(40 citation statements)

References 40 publications

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“…38 These results show how an almost identical DmsAL peptide (DmsAL 1-42 or DmsAL 1-43 ) fused to different partner proteins and assayed at different pH, influence the interaction with DmsD. When smaller portions of the complete DmsAL peptidesequence were assayed for binding to DmsD, the tightest K d (1.7 µM) was observed for the peptide used here (DmsAL 24. When Ser15 and the first arginine (Arg16) were removed from the peptide, binding to DmsD was reduced (K d = 2.1 µM).…”

mentioning

confidence: 76%

“…24 The same DmsA leader peptide sequence ( Figure 1C) was used here to evaluate the binding of each of the 14 single substitution (Table 2).…”

Section: Thermodynamics Of Interaction Between Dmsd Variant Proteins mentioning

confidence: 99%

“…24 The conformational entropy, or the entropy gained/lost within the protein itself (due to reduced/increased freedom) is thought to be small as it is assumed that the protein structure becomes more rigid upon binding its substrate. In fact, upon binding, most peptides do not induce conformational changes in their partner protein, minimizing this entropic cost.…”

Section: Stoichiometry Of Binding and Hydrodynamic Characterization Omentioning

confidence: 99%

“…[24][25][26] A synthetic peptide composed of residues 15 through 41 of the 45 amino acid DmsA leader (DmsAL) peptide was shown to bind to DmsD with micro molar affinity (1.7 µM) and the minimal binding unit of the DmsAL peptide was shown to be the hydrophobic region (DmsAL . 24 A putative RR leader peptide binding site was identified using an immunoblot Far-Western technique to assess binding of DmsD single substitution variant proteins and a modeled structure of DmsD. 27 Subsequently, the structure of the E. coli DmsD (EcDmsD) protein was reported, and amino acids previously shown to be important for binding were found clustered within a hydrophobic pocket of the structure.…”

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confidence: 99%

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Thermodynamic Characterization of the DmsD Binding Site for the DmsA Twin-Arginine Motif

Winstone

Turner

2015

Biochemistry

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The system specific chaperone DmsD interacts with the twin-arginine leader peptide of its substrate, DmsA, allowing for proper folding and assembly of the DmsA catalytic subunit of dimethyl sulfoxide reductase prior to translocation by the twin-arginine translocase. DmsD residues important for binding the complete 45-amino acid sequence of the DmsA leader (DmsAL) peptide were previously identified and found to cluster in a pocket of the DmsD structure. In this study, we have utilized isothermal titration calorimetry (ITC) to determine the dissociation constant and thermodynamic parameters of 15 single-substitution DmsD variant proteins and a synthetic DmsAL peptide consisting of 27 amino acids (DmsAL₁₅₋₄₁). The stoichiometry values were determined via ITC, and the multimeric compositions of the DmsD variants in the absence and presence of peptide were characterized via size exclusion chromatography and native polyacrylamide gel electrophoresis. An up to 4-fold change in affinity was observed for DmsD variant proteins relative to that of wild-type DmsD, and variation of the entropic contribution to binding divided the binding site into two clusters: residues with either more or less favorable entropy. Substitution of hydrophobic residues along one helix face (helix 5) or prolines found on adjacent loops caused reduced binding affinity because of the increased entropic cost, which suggests that the twin-arginine motif of the DmsAL peptide binds to a preformed site on DmsD. Most DmsD variants were more than 90% monomeric in solution and bound a single peptide per protein molecule. The DmsD variant with the largest dimer population showed increased affinity and induced the formation of tetramers in the presence of peptide, suggesting that dimeric DmsD or an alternatively folded form of DmsD may play an as yet undefined role in binding.

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mentioning

confidence: 76%

“…24 The same DmsA leader peptide sequence ( Figure 1C) was used here to evaluate the binding of each of the 14 single substitution (Table 2).…”

Section: Thermodynamics Of Interaction Between Dmsd Variant Proteins mentioning

confidence: 99%

Section: Stoichiometry Of Binding and Hydrodynamic Characterization Omentioning

confidence: 99%

mentioning

confidence: 99%

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Thermodynamic Characterization of the DmsD Binding Site for the DmsA Twin-Arginine Motif

Winstone

Turner

2015

Biochemistry

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“…Twinarginine leader peptide binding is a canonical feature of REMPs, and as such, binding of DmsD to the DmsA RRleader peptide is an absolute prerequisite for DMSO reductase biogenesis (26,27). It is clear that DmsD binds the DmsA RR-leader at 1:1 stoichiometry and that the entire hydrophobic region is required for binding (46) although specificity is more difficult to define (47). Phylogenetic analyses found DmsD to be related to system-specific chaperones of other MobisPGD-containing oxidoreductases that included TorD for TMAO reductase and NarJ for cytoplasmic nitrate reductase A, both of which are CISM enzymes related to DMSO reductase (19,25).…”

Section: So Dmso] (4)mentioning

confidence: 99%

Assembly pathway of a bacterial complex iron sulfur molybdoenzyme

Cherak

Turner

2017

Biomolecular Concepts

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Protein folding and assembly into macromolecule complexes within the living cell are complex processes requiring intimate coordination. The biogenesis of complex iron sulfur molybdoenzymes (CISM) requires use of a system specific chaperone -a redox enzyme maturation protein (REMP) -to help mediate final folding and assembly. The CISM dimethyl sulfoxide (DMSO) reductase is a bacterial oxidoreductase that utilizes DMSO as a final electron acceptor for anaerobic respiration. The REMP DmsD strongly interacts with DMSO reductase to facilitate folding, cofactor-insertion, subunit assembly and targeting of the multi-subunit enzyme prior to membrane translocation and final assembly and maturation into a bioenergetic catalytic unit. In this article, we discuss the biogenesis of DMSO reductase as an example of the participant network for bacterial CISM maturation pathways.

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Applications of isothermal titration calorimetry - the research and technical developments from 2011 to 2015

Falconer

2016

J. Mol. Recognit.

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Isothermal titration calorimetry is a widely used biophysical technique for studying the formation or dissociation of molecular complexes. Over the last 5 years, much work has been published on the interpretation of isothermal titration calorimetry (ITC) data for single binding and multiple binding sites. As over 80% of ITC papers are on macromolecules of biological origin, this interpretation is challenging. Some researchers have attempted to link the thermodynamics constants to events at the molecular level. This review highlights work carried out using binding sites characterized using x‐ray crystallography techniques that allow speculation about individual bond formation and the displacement of individual water molecules during ligand binding and link these events to the thermodynamic constants for binding. The review also considers research conducted with synthetic binding partners where specific binding events like anion‐π and π‐π interactions were studied. The revival of assays that enable both thermodynamic and kinetic information to be collected from ITC data is highlighted. Lastly, published criticism of ITC research from a physical chemistry perspective is appraised and practical advice provided for researchers unfamiliar with thermodynamics and its interpretation. Copyright © 2016 John Wiley & Sons, Ltd.

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The Hydrophobic Region of the DmsA Twin-Arginine Leader Peptide Determines Specificity with Chaperone DmsD

Cited by 16 publications

References 40 publications

Thermodynamic Characterization of the DmsD Binding Site for the DmsA Twin-Arginine Motif

Thermodynamic Characterization of the DmsD Binding Site for the DmsA Twin-Arginine Motif

Assembly pathway of a bacterial complex iron sulfur molybdoenzyme

Applications of isothermal titration calorimetry - the research and technical developments from 2011 to 2015

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