Comparison of the Binding of Cadmium(II), Mercury(II), and Arsenic(III) to the de Novo Designed Peptides TRI L12C and TRI L16C

Matzapetakis, Manolis; Farrer, Brian T.; Weng, Tsu Chien; Hemmingsen, Lars; Penner‐Hahn, James E.; Pecoraro, Vincent L.

doi:10.1021/ja017520u

Cited by 127 publications

(238 citation statements)

References 66 publications

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“…By changing the leucine layer above the metal to alanine to allow better water access above the metalbinding site, we designed a peptide (TRI L12AL16C) that binds Cd(II) solely in a four-coordinate geometry, with three cysteinyl thiolates and one exogenous water. This observation would suggest that Cd(II) binds to these peptides as the exo conformer, consistent with the longer Cd(II)-S distances (2.54 Å), which are more reminiscent of Pb(II) than As(III) (8). These observations make one consider whether metalloregulatory proteins may be able to differentiate different toxic metals in part by their preference for endo [As(III)] versus exo [Pb(II)/Cd(II)] binding.…”

Section: Discussionmentioning

confidence: 56%

“…Previously, we have shown that the three-stranded coiled coil TRI peptides [Ac-G(L a K b A c L d E e E f K g ) 4 G-NH 2 ] designed with the heptad repeat strategy, such as TRI L16C (sequences of all peptides are in Table 1), were able to model the unusual trigonal thiolate Hg(II) coordination geometry seen in the MerR metalloregulatory protein (7). We have also examined the binding of the heavy metals Cd(II), Pb(II), and Bi(III) to the TRI peptides to better understand their modes of toxicity (8,9). Based on this success, we examined whether the trigonal thiolate metal-binding cavities of TRI derivatives would be good models of the active site of ArsR.…”

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confidence: 99%

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Identifying important structural characteristics of arsenic resistance proteins by using designed three-stranded coiled coils

Touw

Nordman

Stuckey³

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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154

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Arsenic, a contaminant of water supplies worldwide, is one of the most toxic inorganic ions. Despite arsenic's health impact, there is relatively little structural detail known about its interactions with proteins. Bacteria such as Escherichia coli have evolved arsenic resistance using the Ars operon that is regulated by ArsR, a repressor protein that dissociates from DNA when As(III) binds. This protein undergoes a critical conformational change upon binding As(III) with three cysteine residues. Unfortunately, structures of ArsR with or without As(III) have not been reported. Alternatively, de novo designed peptides can bind As(III) in an endo configuration within a thiolate-rich environment consistent with that proposed for both ArsR and ArsD. We report the structure of the As(III) complex of Coil Ser L9C to a 1.8-Å resolution, providing x-ray characterization of As(III) in a Tris thiolate protein environment and allowing a structural basis by which to understand arsenated ArsR.arsenic-binding proteins ͉ coiled coil peptides ͉ crystallography ͉ heavy metal toxicity ͉ protein design A rsenic toxicity is a worldwide problem as a natural contaminant of water supplies. Despite the fact that it is a human toxin and carcinogen, few structural reports on its interaction with biological ligands have appeared. Escherichia coli and other bacteria have evolved a detoxification mechanism that employs the arsRDABC operon (1). Arsenic removal by these encoded proteins is initiated when As(III) binds to ArsR, resulting in the dissociation of the repressor protein from the promoter DNA. The structure of the ArsA component of the ATP-dependent extrusion pump ArsAB with antimonite bound in close proximity to the nucleotide-binding site was able to provide some insight into the active transport of As(III) out of the cell (2). Additionally, structural characterization of substrate and product complexes of the arsenate reductase ArsC, which reduces arsenate (AsO 4 3Ϫ ) to arsenite (AsO 2 Ϫ ), has helped elucidate this step of the arsenic detoxification pathway (3, 4). Recent studies of E. coli ArsD indicate that it is a metallochaperone, transporting As(III) to ArsA for extrusion (5). Extended x-ray absorption fine structure and mutagenesis studies have shown that ArsR coordinates As(III) with three cysteine thiolates at a distance of 2.25 Å, a coordination mode that ArsD is also proposed to employ (1, 5). However, no x-ray or NMR structures of ArsR, the repressor protein, or ArsD have been reported to date. Furthermore, AsS 3 structures have not been reported for any biologically relevant small-molecule thiolates, such as glutathione, and only a handful of related AsS 3 complexes with aromatic ligands or chelated alkyl dithiolate coordination are reported (6).We set out to model the putative As(III) coordination environments of ArsR and ArsD in a designed peptide system. Previously, we have shown that the three-stranded coiled coildesigned with the heptad repeat strategy, such as TRI L16C (sequences of all peptides are in Table 1...

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Section: Discussionmentioning

confidence: 56%

mentioning

confidence: 99%

Identifying important structural characteristics of arsenic resistance proteins by using designed three-stranded coiled coils

Touw

Nordman

Stuckey³

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

154

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“…PAC Spectroscopy. PAC spectroscopy and sample preparation were performed as described (35,36); however, a typing error of ''10 -40 mL'' should be ''10 -40 L,'' and the current experiment with 111m Cd and TRIL12LDL16C was carried out in a 1:12 ratio, at pH 9.1, 1°C, and in 55% wt/wt sucrose to further reduce the Brownian tumbling of the molecules. Finally, all fits were carried out with 300 data points, disregarding the 5 first points due to systematic errors in these.…”

Section: Methodsmentioning

confidence: 99%

Using diastereopeptides to control metal ion coordination in proteins

Peacock

Hemmingsen

Pecoraro

2008

Proc. Natl. Acad. Sci. U.S.A.

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Here, we report a previously undescribed approach for controlling metal ion coordination geometry in biomolecules by reorientating amino acid side chains through substitution of L-to D-amino acids. These diastereopeptides allow us to manipulate the spatial orientation of amino acid side chains to alter the sterics of metal binding pockets. We have used this approach to design the de novo metallopeptide, Cd (TRIL12L DL16C)

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“…The simplest way is to analyze protein scaffolds empirically and introduce cysteine-rich coordination sites. Several α-helical coiled-coil metalloproteins capable of binding Co(II), Fe(II), Zn(II), Cd(II), Hg(II), and As(II) have been designed this way (Matzapetakis et al, 2002;Petros et al, 2006). More complex coordination sites were engineered with the aid of computational searching, for example, the Cys 2 -His 2 zinc binding sites and Fe 4 -S 4 sites (Lu et al, 2001).…”

Section: Discussionmentioning

confidence: 99%

Engineering a zinc binding site into the de novo designed protein DS119 with a βαβ structure

Zhu¹,

Liang²,

Lai³

2011

Protein Cell

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Functional proteins designed de novo have potential application in chemical engineering, agriculture and healthcare. Metal binding sites are commonly used to incorporate functions. Based on a de novo designed protein DS119 with a βαβ structure, we have computationally engineered zinc binding sites into it using a home-made searching program. Seven out of the eight designed sequences tested were shown to bind Zn 2+ with micromolar affinity, and one of them bound Zn 2+ with 1:1 stoichiometry. This is the first time that metalloproteins with an α, β mixed structure have been designed from scratch.

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Comparison of the Binding of Cadmium(II), Mercury(II), and Arsenic(III) to the de Novo Designed Peptides TRI L12C and TRI L16C

Cited by 127 publications

References 66 publications

Identifying important structural characteristics of arsenic resistance proteins by using designed three-stranded coiled coils

Identifying important structural characteristics of arsenic resistance proteins by using designed three-stranded coiled coils

Using diastereopeptides to control metal ion coordination in proteins

Engineering a zinc binding site into the de novo designed protein DS119 with a βαβ structure

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