Controlling Protein−Protein Interactions through Metal Coordination:  Assembly of a 16-Helix Bundle Protein

Salgado, Eric N.; Faraone-Mennella, Jasmin; Tezcan, F.A.

doi:10.1021/ja075261o

Cited by 125 publications

(192 citation statements)

References 17 publications

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“…1B, Species 2 in Fig. 1A) (7,8). Such i, i þ 4 diHis motifs have been used for the assembly and stabilization of metalloproteins and peptides through coordination of divalent transition metal ions (9)(10)(11)(12).…”

Section: Resultsmentioning

confidence: 99%

“…S3). Previous SV measurements on MBPC-1 indicated that the predominant species in solution at low protein and equimolar Zn concentrations was monomeric (7). Dimeric and tetrameric species become significantly populated only at MBPC-1 and Zn concentrations over 100 μM (1∶1 protein∶Zn) with increasing concentrations favoring the population of the tetrameric form.…”

Section: Redesign Of I2mentioning

confidence: 94%

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Metal templated design of protein interfaces

Salgado

Ambroggio

Brodin

et al. 2009

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

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130

156

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Metal coordination is a key structural and functional component of a large fraction of proteins. Given this dual role we considered the possibility that metal coordination may have played a templating role in the early evolution of protein folds and complexes. We describe here a rational design approach, Metal Templated Interface Redesign (MeTIR), that mimics the time course of a hypothetical evolutionary pathway for the formation of stable protein assemblies through an initial metal coordination event. Using a folded monomeric protein, cytochrome cb 562 , as a building block we show that its non-self-associating surface can be made self-associating through a minimal number of mutations that enable Zn coordination. The protein interfaces in the resulting Zn-directed, D 2 -symmetrical tetramer are subsequently redesigned, yielding unique protein architectures that self-assemble in the presence or absence of metals. Aside from its evolutionary implications, MeTIR provides a route to engineer de novo protein interfaces and metal coordination environments that can be tuned through the extensive noncovalent bonding interactions in these interfaces.evolution | metal coordination | protein self-assembly | protein-protein interactions | templating

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Section: Resultsmentioning

confidence: 99%

Section: Redesign Of I2mentioning

confidence: 94%

Metal templated design of protein interfaces

Salgado

Ambroggio

Brodin

et al. 2009

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

Self Cite

130

156

View full text Add to dashboard Cite

show abstract

“…Recently, the Tezcan (11) and Kuhlman (12) groups used rational and computational protein design methods, respectively, to engineer metal-dependent protein-protein interactions using conserved dihistidine metal-binding motifs from known metalloproteins. In each case, structural analysis of the engineered protein complexes suggested that, although metal-dependent assembly had been achieved, the metalbinding sites had not formed as desired (11,12). Although notable successes, these studies highlight the difficulty in precisely stabilizing the conformations of multiple amino acid side chains at once.…”

mentioning

confidence: 99%

Computational design of a homotrimeric metalloprotein with a trisbipyridyl core

Mills

Sheffler

Ener

et al. 2016

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

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Metal-chelating heteroaryl small molecules have found widespread use as building blocks for coordination-driven, self-assembling nanostructures. The metal-chelating noncanonical amino acid (2,2′-bipyridin5yl)alanine (Bpy-ala) could, in principle, be used to nucleate specific metalloprotein assemblies if introduced into proteins such that one assembly had much lower free energy than all alternatives. Here we describe the use of the Rosetta computational methodology to design a self-assembling homotrimeric protein with [Fe (Bpy-ala) 3 ] 2+ complexes at the interface between monomers. X-ray crystallographic analysis of the homotrimer showed that the design process had near-atomic-level accuracy: The all-atom rmsd between the design model and crystal structure for the residues at the protein interface is ∼1.4 Å. These results demonstrate that computational protein design together with genetically encoded noncanonical amino acids can be used to drive formation of precisely specified metal-mediated protein assemblies that could find use in a wide range of photophysical applications.computational protein design | noncanonical amino acids | metalloproteins | protein self-assembly T he small-molecule metal ligand 2,2′-bipyridine (Bpy) has found widespread use in inorganic chemistry because of its redox stability, ability to form high-affinity complexes of defined geometry with a variety of transition metals, and useful photochemical properties (1). The highly specific geometries adopted by Bpy in metal complexes have been used to generate coordinationdriven self-assembling nanostructures with defined structures (2). For example, chemically synthesized, unstructured peptides containing the Bpy functional group as the side chain of an amino acid formed three-helix bundles upon addition of metals (3, 4). Recently, the ability to use Bpy in biological contexts was expanded through the addition of the noncanonical amino acid (NCAA) (2,2′-bipyridin-5yl)alanine (Bpy-ala) to the genome of Escherichia coli (5 2+ complexes (where M is a divalent cation that forms an octahedral complex with Bpy). Although we (6) and others (7-9) have engineered proteins in which Bpy-ala served in structural or functional capacities, to our knowledge, the possibility of using this NCAA to drive or stabilize the formation of a protein complex has not been explored.The strategy of using metal ions to mediate protein-complex formation has its origins in naturally occurring proteins, such as hexameric proinsulin, in which complex formation is regulated by bound Ca 2+ and Zn 2+ ions (10). Recently, the Tezcan (11) and Kuhlman (12) groups used rational and computational protein design methods, respectively, to engineer metal-dependent protein-protein interactions using conserved dihistidine metal-binding motifs from known metalloproteins. In each case, structural analysis of the engineered protein complexes suggested that, although metal-dependent assembly had been achieved, the metalbinding sites had not formed as desired (11,12). Although notable su...

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“…[28,29] The resulting products favored dimers and tetramers often as mixtures, which could be tailored to favor a specific oligomer. Other groups have also employed various metal ion binding motifs to stabilize a designed protein motif; however, few are designed with the specific intent of controlling the oligomeric state.…”

mentioning

confidence: 99%

Conformationally Constrained Sequence Designs to Bias Monomer–Dimer Equilibriums in TASP Systems

Freeman

Sherman

2011

Chemistry A European J

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We have designed template-assembled synthetic proteins (TASPs) with the intent of controlling their oligomeric state by stabilizing specific helical tertiary structures via histidine metal ion chelation or disulfide incorporation. In solution, cavitein Q4 was previously determined to interconvert between a four-helix bundle monomer and an eight-helix bundle dimer. In this paper, we show that judicious mutation of cavitein Q4 can stabilize either the monomeric parallel four-helix bundle or the dimeric antiparallel eight-helix bundle structure. Cavitein Q4-E3H, designed to be dimeric, is indeed biased toward dimerization as a result of incorporation of histidines. Moreover, the addition of nickel was found to further increase the association constant of dimerization. Similarly, a cavitein designed to stabilize the monomeric structure via histidine metal ion chelation (Q4-H) was found to favor a monomer in solution upon addition of nickel. Lastly, a cavitein intended to stabilize a monomeric structure via disulfide incorporation (Q4-C2) is reported. Surprisingly, this disulfide cavitein yielded two products upon oxidation suggesting disulfide formation both above the cavitand template and below may be possible. Nevertheless, the two disulfide caviteins were shown to exist as monomers as per their design.

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Controlling Protein−Protein Interactions through Metal Coordination: Assembly of a 16-Helix Bundle Protein

Cited by 125 publications

References 17 publications

Metal templated design of protein interfaces

Metal templated design of protein interfaces

Computational design of a homotrimeric metalloprotein with a trisbipyridyl core

Conformationally Constrained Sequence Designs to Bias Monomer–Dimer Equilibriums in TASP Systems

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