Spectroscopic and metal binding properties of a <i>de novo</i> metalloprotein binding a tetrazinc cluster

Chino, Marco; Zhang, Shao-Qing; Pirro, Fabio; Leone, Linda; Maglio, Ornella; Lombardi, Angela; DeGrado, William F.

doi:10.1002/bip.23229

Cited by 17 publications

(11 citation statements)

References 49 publications

(55 reference statements)

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“…Most recently, the design principles used in the construction of DF proteins have recently been extended to engineer tetranuclear Zn 2+ clusters (Chino et al ., 2018; Zhang et al ., 2018 a ). The site included four bridging Asp and four terminal His ligands, as well as a total of 16 polar side chains in a fully connected hydrogen-bonded network (Fig.…”

Section: Computational Design Guided By Fundamental Physicochemical Pmentioning

confidence: 99%

De novoprotein design, a retrospective

Korendovych

DeGrado

2020

Quart. Rev. Biophys.

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176

179

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Proteins are molecular machines whose function depends on their ability to achieve complex folds with precisely defined structural and dynamic properties. The rational design of proteins from first-principles, or de novo, was once considered to be impossible, but today proteins with a variety of folds and functions have been realized. We review the evolution of the field from its earliest days, placing particular emphasis on how this endeavor has illuminated our understanding of the principles underlying the folding and function of natural proteins, and is informing the design of macromolecules with unprecedented structures and properties. An initial set of milestones in de novo protein design focused on the construction of sequences that folded in water and membranes to adopt folded conformations. The first proteins were designed from first-principles using very simple physical models. As computers became more powerful, the use of the rotamer approximation allowed one to discover amino acid sequences that stabilize the desired fold. As the crystallographic database of protein structures expanded in subsequent years, it became possible to construct proteins by assembling short backbone fragments that frequently recur in Nature. The second set of milestones in de novo design involves the discovery of complex functions. Proteins have been designed to bind a variety of metals, porphyrins, and other cofactors. The design of proteins that catalyze hydrolysis and oxygen-dependent reactions has progressed significantly. However, de novo design of catalysts for energetically demanding reactions, or even proteins that bind with high affinity and specificity to highly functionalized complex polar molecules remains an importnant challenge that is now being achieved. Finally, the protein design contributed significantly to our understanding of membrane protein folding and transport of ions across membranes. The area of membrane protein design, or more generally of biomimetic polymers that function in mixed or non-aqueous environments, is now becoming increasingly possible.

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Section: Computational Design Guided By Fundamental Physicochemical Pmentioning

confidence: 99%

De novoprotein design, a retrospective

Korendovych

DeGrado

2020

Quart. Rev. Biophys.

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176

179

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“…Second, the structure of the designed multicofactor protein was determined by X-ray crystallography. It is also noteworthy that the computational methods, adopted in this work, could be readily extended to the design of electronically coupled systems for light-triggered electron energy storage and utilization, particularly given the ability to design proteins that incorporate metal ion clusters ( 15 , 21 , 23 – 31 ).…”

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confidence: 99%

Allosteric cooperation in a de novo-designed two-domain protein

Pirro

Schmidt

Lincoff

et al. 2020

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

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We describe the de novo design of an allosterically regulated protein, which comprises two tightly coupled domains. One domain is based on the DF (Due Ferri in Italian or two-iron in English) family of de novo proteins, which have a diiron cofactor that catalyzes a phenol oxidase reaction, while the second domain is based on PS1 (Porphyrin-binding Sequence), which binds a synthetic Zn-porphyrin (ZnP). The binding of ZnP to the original PS1 protein induces changes in structure and dynamics, which we expected to influence the catalytic rate of a fused DF domain when appropriately coupled. Both DF and PS1 are four-helix bundles, but they have distinct bundle architectures. To achieve tight coupling between the domains, they were connected by four helical linkers using a computational method to discover the most designable connections capable of spanning the two architectures. The resulting protein, DFP1 (Due Ferri Porphyrin), bound the two cofactors in the expected manner. The crystal structure of fully reconstituted DFP1 was also in excellent agreement with the design, and it showed the ZnP cofactor bound over 12 Å from the dimetal center. Next, a substrate-binding cleft leading to the diiron center was introduced into DFP1. The resulting protein acts as an allosterically modulated phenol oxidase. Its Michaelis–Menten parameters were strongly affected by the binding of ZnP, resulting in a fourfold tighter Km and a 7-fold decrease in kcat. These studies establish the feasibility of designing allosterically regulated catalytic proteins, entirely from scratch.

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“…Zhang and Lombardi et al have taken another step closer toward reproducing a full Mn cluster seen in PSII by designing a protein capable of binding a tetra-zinc cluster [ 27 , 28 ]. Their design uses the four Zn atoms to form anchor points for four separate helices resulting in a homotetrameric assembly.…”

Section: Small Molecule Binding Proteinsmentioning

confidence: 99%

Recent Progress Using De Novo Design to Study Protein Structure, Design and Binding Interactions

Ferrando

Solomon

2021

Life

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De novo protein design is a powerful methodology used to study natural functions in an artificial-protein context. Since its inception, it has been used to reproduce a plethora of reactions and uncover biophysical principles that are often difficult to extract from direct studies of natural proteins. Natural proteins are capable of assuming a variety of different structures and subsequently binding ligands at impressively high levels of both specificity and affinity. Here, we will review recent examples of de novo design studies on binding reactions for small molecules, nucleic acids, and the formation of protein-protein interactions. We will then discuss some new structural advances in the field. Finally, we will discuss some advancements in computational modeling and design approaches and provide an overview of some modern algorithmic tools being used to design these proteins.

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Spectroscopic and metal binding properties of a de novo metalloprotein binding a tetrazinc cluster

Cited by 17 publications

References 49 publications

De novoprotein design, a retrospective

De novoprotein design, a retrospective

Allosteric cooperation in a de novo-designed two-domain protein

Recent Progress Using De Novo Design to Study Protein Structure, Design and Binding Interactions

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