Tryptophan-scanning mutagenesis of the ligand binding pocket in Thermotoga maritima arginine-binding protein

Deacon, Lindsay J.; Billones, Hilbert; Galyean, Anne A.; Donaldson, Teraya; Pennacchio, Anna; Iozzino, Luisa; D’Auria, Sabato; Dattelbaum, Jonathan D.

doi:10.1016/j.biochi.2013.12.011

Cited by 11 publications

(10 citation statements)

References 38 publications

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“…In line with previous reports [7], [9], gel filtration analysis indicates that the protein adopts different aggregation states (data not shown). Although aggregates with higher molecular masses were detected, we focused our attention on the main component.…”

Section: Resultssupporting

confidence: 93%

“…The protein binds the substrate through an intricate network of interactions, which results in a high affinity of binding at room temperature (K D in the nanomolar range). Although this may appear in contrast with previous reports [7], the analysis of the 3D structure reconciles all data. Indeed, SPR experiments indicated a much lower affinity (K D of 17 µM) of the protein for a rather complex arginine-containing peptide [9].…”

Section: Discussioncontrasting

confidence: 50%

“…In this case, however, tertiary structure variations associated with ligand binding are very limited. The oligomerization through domain swapping exhibited by TmArgBP also provides a rationale for the observed ability of the protein to form higher aggregates [7], [9]. These states may be achieved through a mutual swapping of the C-terminal helix that is not limited to two subunits but involves three (trimers) or four (tetramers) protein molecules, as found in RNase A [13], [21], [22].…”

Section: Discussionmentioning

confidence: 99%

“…We identified the arginine binding protein form Thermatoga maritima (TmArgBP) as an ideal system for arginine detection [6], [7], [8], [9], [10]. Arginine sensing is extremely important since argininemia is a debilitating inherited condition, characterized by a gradual accumulation of arginine and ammonia in the blood, whose diagnosis is crucial for effective medical intervention [11].…”

Section: Introductionmentioning

confidence: 99%

“…These include an extraordinary stability to both temperature and chemical denaturants [9]. Additionally, in contrast to the vast majority of SBP that operate as monomers, TmArgBP forms multimeric assemblies at room temperature [7], [9].…”

Section: Introductionmentioning

confidence: 99%

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A Loose Domain Swapping Organization Confers a Remarkable Stability to the Dimeric Structure of the Arginine Binding Protein from Thermotoga maritima

Ruggiero

Dattelbaum

Staiano³

et al. 2014

PLoS ONE

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The arginine binding protein from Thermatoga maritima (TmArgBP), a substrate binding protein (SBP) involved in the ABC system of solute transport, presents a number of remarkable properties. These include an extraordinary stability to temperature and chemical denaturants and the tendency to form multimeric structures, an uncommon feature among SBPs involved in solute transport. Here we report a biophysical and structural characterization of the TmArgBP dimer. Our data indicate that the dimer of the protein is endowed with a remarkable stability since its full dissociation requires high temperature as well as SDS and urea at high concentrations. In order to elucidate the atomic level structural properties of this intriguing protein, we determined the crystallographic structures of the apo and the arginine-bound forms of TmArgBP using MAD and SAD methods, respectively. The comparison of the liganded and unliganded models demonstrates that TmArgBP tertiary structure undergoes a very large structural re-organization upon arginine binding. This transition follows the Venus Fly-trap mechanism, although the entity of the re-organization observed in TmArgBP is larger than that observed in homologous proteins. Intriguingly, TmArgBP dimerizes through the swapping of the C-terminal helix. This dimer is stabilized exclusively by the interactions established by the swapping helix. Therefore, the TmArgBP dimer combines a high level of stability and conformational freedom. The structure of the TmArgBP dimer represents an uncommon example of large tertiary structure variations amplified at quaternary structure level by domain swapping. Although the biological relevance of the dimer needs further assessments, molecular modelling suggests that the two TmArgBP subunits may simultaneously interact with two distinct ABC transporters. Moreover, the present protein structures provide some clues about the determinants of the extraordinary stability of the biomolecule. The availability of an accurate 3D model represents a powerful tool for the design of new TmArgBP suited for biotechnological applications.

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Section: Resultssupporting

confidence: 93%

Section: Discussioncontrasting

confidence: 50%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

A Loose Domain Swapping Organization Confers a Remarkable Stability to the Dimeric Structure of the Arginine Binding Protein from Thermotoga maritima

Ruggiero

Dattelbaum

Staiano³

et al. 2014

PLoS ONE

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Tryptophan Scanning Mutagenesis of EF-Hand Motifs

Kiran

Kreutz

Sharma

et al. 2019

Methods in Molecular Biology

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Studies of conformational changes of an arginine-binding protein from Thermotoga maritima in the presence and absence of ligand via molecular dynamics simulations with the coarse-grained UNRES force field

et al. 2015

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The arginine-binding protein (ArgBP) from the hyperthermophilic eubacterium Thermotoga maritima (TmArgBP) is responsible for arginine transport through the bacterial cell membrane. The protein binds a single molecule of L-arginine, which results in conformational changes due to hinge bending. Thereby, TmArgBP acquires one of two possible conformations: open (without the presence of the arginine ligand) and closed (in the presence of the arginine ligand). Here we report a molecular dynamics study of the influence of the presence or absence of the ligand on the dynamics of TmArgBP, using the coarse-grained UNRES force field. The results of our studies indicate that binding of the arginine ligand promotes a closed conformation, which agrees with experimental data. However, the sensitivity of the TmArgBP conformation to the presence of arginine decreases and the protein becomes more flexible with increasing temperature, which might be related to the functionality of this protein in the thermophilic organism T. maritima.

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Tryptophan-scanning mutagenesis of the ligand binding pocket in Thermotoga maritima arginine-binding protein

Cited by 11 publications

References 38 publications

A Loose Domain Swapping Organization Confers a Remarkable Stability to the Dimeric Structure of the Arginine Binding Protein from Thermotoga maritima

A Loose Domain Swapping Organization Confers a Remarkable Stability to the Dimeric Structure of the Arginine Binding Protein from Thermotoga maritima

Tryptophan Scanning Mutagenesis of EF-Hand Motifs

Studies of conformational changes of an arginine-binding protein from Thermotoga maritima in the presence and absence of ligand via molecular dynamics simulations with the coarse-grained UNRES force field

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