Dataset Information

Photocatalytic Hydrogen Evolution by a De Novo Designed Metalloprotein that Undergoes Ni-Mediated Oligomerization Shift.

ABSTRACT: De novo metalloprotein design involves the construction of proteins guided by specific repeat patterns of polar and apolar residues, which, upon self-assembly, provide a suitable environment to bind metals and produce artificial metalloenzymes. While a wide range of functionalities have been realized in de novo designed metalloproteins, the functional repertoire of such constructs towards alternative energy-relevant catalysis is currently limited. Here we show the application of de novo approach to design a functional H₂ evolving protein. The design involved the assembly of an amphiphilic peptide featuring cysteines at tandem a/d sites of each helix. Intriguingly, upon Ni^II addition, the oligomers shift from a major trimeric assembly to a mix of dimers and trimers. The metalloprotein produced H₂ photocatalytically with a bell-shape pH dependence, having a maximum activity at pH 5.5. Transient absorption spectroscopy is used to determine the timescales of electron transfer as a function of pH. Selective outer sphere mutations are made to probe how the local environment tunes activity. A preferential enhancement of activity is observed via steric modulation above the Ni^II site, towards the N-termini, compared to below the Ni^II site towards the C-termini.

SUBMITTER: Prasad P

PROVIDER: S-EPMC10308963 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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Publications

Photocatalytic Hydrogen Evolution by a De Novo Designed Metalloprotein that Undergoes Ni-Mediated Oligomerization Shift.

Prasad Pallavi P Hunt Leigh Anna LA Pall Ashley E AE Ranasinghe Maduni M Williams Ashley E AE Stemmler Timothy L TL Demeler Borries B Hammer Nathan I NI Chakraborty Saumen S

Chemistry (Weinheim an der Bergstrasse, Germany) 20230206 14

De novo metalloprotein design involves the construction of proteins guided by specific repeat patterns of polar and apolar residues, which, upon self-assembly, provide a suitable environment to bind metals and produce artificial metalloenzymes. While a wide range of functionalities have been realized in de novo designed metalloproteins, the functional repertoire of such constructs towards alternative energy-relevant catalysis is currently limited. Here we show the application of de novo approach ...[more]

PMID: 36440875

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Project description:The relationship between structure and function has long been one of the major points of investigation in Biophysics. Understanding how much, or how little, of a protein's often complicated structure is necessary for its function can lead to directed therapeutic strategies and would allow one to design proteins for specific desired functions. Studying protein function by de novo design builds the functionality from the ground up in a completely unrelated and noncoded protein scaffold. Our lab has used this strategy to study heavy and transition metal binding within the TRI family of three stranded coiled coil (3SCC) constructs to understand coordination geometry and metalloenzyme catalytic control within a protein environment. These peptides contain hydrophobic layers within the interior of the 3SCC, which one can mutate to metal binding residues to create a minimal metal binding site, while solid phase synthesis allows our lab to easily incorporate a number of noncoded amino acids including d enantiomers of binding or secondary coordination sphere amino acids, penicillamine, or methylated versions of histidine. Our studies of Cd(II) binding to Cys3 environments have determined, largely through the use of 113Cd NMR and 111mCd PAC, that the coordination environment around a heavy metal can be controlled by incorporating noncoded amino acids in either the primary or secondary coordination spheres. We found mutating the metal binding amino acids to l-Pen can enforce trigonal Cd(II)S3 geometry exclusively compared to the mixed coordination determined for l-Cys coordination. The same result can be achieved with secondary sphere mutations as well by incorporating d-Leu above a Cys3. We hypothesize this latter effect is due to the increased steric packing above the metal binding site that occurs when the l-Leu oriented toward the N-terminus of the scaffold is mutated to d-Leu and oriented toward the C-terminus. Mutating the layer below Cys3 to d-Leu instead formed a mixed 4- and 5-coordinate Cd(II)S3(H2O) and Cd(II)S3(H2O)2 construct as steric bulk was decreased below the metal binding site. We have also applied noncoded amino acids to metalloenzyme systems by incorporating His residues that are methylated at the δ- or ε-nitrogen to enforce Cu(I) ligation to the opposite open nitrogen of His and found a 2 orders of magnitude increased catalytic efficiency for nitrite reductase activity with ε-nitrogen coordination compared to δ-nitrogen. These results exemplify the ability to tune coordination environment and catalytic efficiency within a de novo scaffold as well as the utility of noncoded amino acids to increase the chemist's toolbox. By furthering our understanding of metalloprotein design one could envision, through our use of amino acids not normally available to nature, that protein design laboratories will soon be capable of outperforming the native systems previously used as their benchmark of successful design. The ability to design proteins at this level would have far reaching and exciting benefits within various fields including medical and industrial applications.

Dataset Information

Photocatalytic Hydrogen Evolution by a De Novo Designed Metalloprotein that Undergoes Ni-Mediated Oligomerization Shift.

Publications

Photocatalytic Hydrogen Evolution by a De Novo Designed Metalloprotein that Undergoes Ni-Mediated Oligomerization Shift.

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