Article Chemistry, Multidisciplinary
A De Novo-Designed Type 3 Copper Protein Tunes Catechol Substrate Recognition and Reactivity
Fabio Pirro, Salvatore La Gatta, Federica Arrigoni, Antonino Famulari, Ornella Maglio, Pompea Del Vecchio, Mario Chiesa, Luca De Gioia, Luca Bertini, Marco Chino, Flavia Nastri, Angela Lombardi
Summary: De novo metalloprotein design is a remarkable method for shaping protein structures to achieve specific functions. In this study, a protein called DR1 was designed and characterized to harbor a di-copper site, mimicking copper-containing polyphenol oxidases. The di-copper site was engineered into a four-helix bundle scaffold through hierarchical design of metal coordination spheres. Spectroscopic, thermodynamic, and functional analyses demonstrated that DR1 exhibited the characteristics of a Type 3 copper site, supporting different copper redox states and catalyzing the oxygen-dependent oxidation of catechols. Careful design of substrate access site residues endowed DR1 with substrate recognition capability.
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION (2023)
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Article Biochemical Research Methods
Effects of Hydrophobic Residues on the Intracellular Self-Assembly of De Novo Designed Peptide Tags and Their Orthogonality
Takayuki Miki, Keigo Kajiwara, Sae Nakayama, Masahiro Hashimoto, Hisakazu Mihara
Summary: Protein assemblies play vital roles in living systems, and self-assembling peptides (SAPs) serve as potential tags. In this study, we investigated the role of hydrophobic residues in SAPs and found that different hydrophobic amino acids significantly affect self-assembly propensity, assembly size, protein denaturation, and subcellular localization. Additionally, we explored the orthogonality of their interactions.
ACS SYNTHETIC BIOLOGY (2022)
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Article Chemistry, Multidisciplinary
A De Novo-Designed Artificial Metallopeptide Hydrogenase: Insights into Photochemical Processes and the Role of Protonated Cys
Sreya Malayam Parambath, Ashley E. Williams, Leigh Anna Hunt, Dhanashree Selvan, Nathan Hammer, Saumen Chakraborty
Summary: This study reports the construction of a de novo-designed artificial hydrogenase (ArH) inspired by [NiFe] hydrogenases, which produces H-2 gas photocatalytically and shows bell-shaped pH-dependence on activity. Spectroscopic studies elucidate the reaction mechanism and a fine balance is found between solution acidity and electron transfer steps for maximizing H-2 production.
CHEMSUSCHEM (2021)
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Article Chemistry, Physical
de Novo-designed antimicrobial peptides with broad-spectrum antimicrobial potency and rapid wound disinfection
Yong Fang Zheng, Shi Xian Chen, Ke Jing Mao, Xin Yu Zhu, Ming Yi Jiang, Chang Jer Wu, Jian Ren Lu, Hu Zhu
Summary: In this study, a short peptide G(IIKK)3I was modified to optimize its antibacterial performance by altering its physicochemical parameters. The results showed that peptides with stronger hydrophobicity exhibited stronger antibacterial activity and faster killing kinetics, but also higher cytotoxicity and hemolytic activity. Among all the designed peptides, Trp-substituting variants GW1 and GW2 showed the strongest antibacterial activity and good selectivity and resistance. Furthermore, changing their chirality from L to D-configuration had no obvious effects on their antibacterial activity and improved their biostability in serum. In vivo experiments confirmed the excellent antibacterial performance and wound healing properties of D-GW1.
COLLOIDS AND SURFACES A-PHYSICOCHEMICAL AND ENGINEERING ASPECTS (2023)
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Review Chemistry, Multidisciplinary
Catalysis and Electron Transfer in De Novo Designed Metalloproteins
Karl J. Koebke, Tyler B. J. Pinter, Winston C. Pitts, Vincent L. Pecoraro
Summary: This review highlights the progress and milestone achievements in the field of de novo metalloprotein design in the past decade, with a focus on heme binding proteins, catalytic sites, and electron transfer sites. The significant contributions to our understanding of these subfields or de novo metalloprotein design are categorized and discussed in the context of history and science. Suggestions for future research directions are also provided.
CHEMICAL REVIEWS (2022)
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Review Chemistry, Multidisciplinary
Catalysis and Electron Transfer in De Novo Designed Metalloproteins
Karl J. Koebke, Tyler B. J. Pinter, Winston C. Pitts, Vincent L. Pecoraro
Summary: This review summarizes the progress and milestone achievements in the field of de novo metalloprotein design in the past decade, with a special focus on new designs within common subfields of bioinorganic study. The reports discussed provide important contributions to our understanding of these subfields and de novo metalloprotein design. The review also offers general suggestions for future directions to advance our understanding and accelerate discovery.
CHEMICAL REVIEWS (2022)
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Article Chemistry, Analytical
Highly Robust de Novo Full-Length Protein Sequencing
Zhi-Biao Mai, Zhong-Hua Zhou, Qing-Yu He, Gong Zhang
Summary: This article introduces a contig-scaffolding strategy for high robustness and accuracy in protein sequence assembly. The strategy minimizes bias in the hydrolysis process by integrating multiple unspecific hydrolysis methods and uses a multistep assembly algorithm with error correction. Experimental results demonstrate the effectiveness of this strategy in assembling protein sequences with high coverage and accuracy, even for membrane proteins.
ANALYTICAL CHEMISTRY (2022)
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Article Materials Science, Multidisciplinary
De novo functional groups designed to enhance neuronal integrin α5β1 binding using deep reinforcement learning
Isuru S. Herath, Jingjie Yeo
Summary: When functionalized, tissue engineering scaffolds like silk fibroin can be used to create implantable devices for nerve tissue regeneration. By using deep reinforcement learning, we were able to design a small-molecule ligand with improved binding energy for integrin a5 beta 1, which is essential for creating devices that aid in nerve tissue repair.
MRS BULLETIN (2023)
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Article Biology
De novo-designed transmembrane domains tune engineered receptor functions
Assaf Elazar, Nicholas J. Chandler, Ashleigh S. Davey, Jonathan Y. Weinstein, Julie Nguyen, Raphael Trenker, Ryan S. Cross, Misty R. Jenkins, Melissa J. Call, Matthew E. Call, Sarel J. Fleishman
Summary: De novo-designed receptor transmembrane domains (TMDs) offer precise control over cellular receptor functions and can be used to program specific oligomeric interactions into chimeric antigen receptors (CARs), resulting in tunable cytokine release and antitumor activity of CAR T cells. This precise design approach provides a new way to engineer receptor structure and activity in synthetic biology.
ELIFE (2022)
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Article Biochemistry & Molecular Biology
Expanding the versatility of natural and de novo designed coiled coils and helical bundles
Mohammad ElGamacy, Birte Hernandez Alvarez
Summary: Natural helical bundles (HBs) are protein folds built of α helices that adopt diverse topologies and functions, ranging from structural scaffolds to molecular switches; Symmetric HBs, such as coiled coils, serve as ideal model systems for studying protein folding and design.
CURRENT OPINION IN STRUCTURAL BIOLOGY (2021)
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Article Multidisciplinary Sciences
De novo designed ice- binding proteins from twist- constrained helices
Robbert J. de Haas, Roderick P. Tas, Danielle van den Broek, Chuanbao Zheng, Hannah Nguyen, Alex Kang, Asim K. Bera, Neil P. King, Ilja K. Voets, Renko de Vries
Summary: Attaining molecular level control over solidification processes is crucial for materials science. Reverse engineering using de novo computational protein design can help understand the structure-activity relationships of ice-binding proteins (IBPs). The study tested the hypothesis that an alpha-helical winter flounder antifreeze protein uses an unusual undertwisting to align its ice-binding threonine residues in the same direction. Results confirmed the hypothesis and suggest possibilities for the computational design of IBPs.
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA (2023)
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Article Multidisciplinary Sciences
De novo designed protein inhibitors of amyloid aggregation and seeding
Kevin A. Murray, Carolyn J. Hu, Sarah L. Griner, Hope Pan, Jeannette T. Bowler, Romany Abskharon, Gregory M. Rosenberg, Xinyi Cheng, Paul M. Seidler, David S. Eisenberg
Summary: Neurodegenerative diseases are characterized by the accumulation of aggregated proteins, and inhibiting the formation of these aggregates is a potential therapeutic strategy. Using de novo protein design, researchers have developed a library of mini-protein inhibitors that specifically target the amyloid structures of tau, Aβ, and α Syn. These inhibitors show promising results in preventing aggregation and rescuing motor deficits in animal models of PD and AD.
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA (2022)
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Review Infectious Diseases
Designed Multifunctional Peptides for Intracellular Targets
Davor Juretic
Summary: This review focuses on peptides that can easily reach intracellular targets with little or no toxicity to mammalian cells, specifically optimized for cell-penetrating, antimicrobial, anticancer, antiviral, antifungal, and anti-inflammatory activity. The review suggests exploring wide-spectrum multifunctionality for novel nontoxic hybrids with cell-penetrating peptides.
ANTIBIOTICS-BASEL (2022)
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Article Biochemistry & Molecular Biology
Substrate promiscuity of a de novo designed peroxidase
Jonathan M. X. Jenkins, Claire E. M. Noble, Katie J. Grayson, Adrian J. Mulholland, J. L. Ross Anderson
Summary: The de novo enzyme C45 exhibits extensive catalytic promiscuity, with substrate activity primarily determined by the redox potential of the substrate and the active oxidising species in peroxidase chemistry. Substrate:protein interactions play a significant role in determining electron transfer rates from substrate to heme, affecting the kinetic parameters of the enzyme. Biomolecular simulation identified potential interactions and binding sites near the heme cofactor, providing insight into the catalytic workings of C45.
JOURNAL OF INORGANIC BIOCHEMISTRY (2021)
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Article Biochemistry & Molecular Biology
De novo designed peptides for cellular delivery and subcellular localisation
Guto G. Rhys, Jessica A. Cross, William M. Dawson, Harry F. Thompson, Sooruban Shanmugaratnam, Nigel J. Savery, Mark P. Dodding, Birte Hoecker, Derek N. Woolfson
Summary: This article describes a de novo synthetic peptide system that can efficiently deliver into cells and has the ability of subcellular targeting.
NATURE CHEMICAL BIOLOGY (2022)
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Article Biochemistry & Molecular Biology
Short self-assembling peptides with a urea bond: A new type of supramolecular peptide hydrogel materials
Hiroshi Tsutsumi, Kunifumi Tanaka, Jyh Yea Chia, Hisakazu Mihara
Summary: Short self-assembling peptides with a symmetric structure via a urea bond were designed and synthesized for hydrogel formation. Both (FFiO)(2) and (FFiK)(2) peptides assembled into beta-sheet structure and formed stable hydrogels, with (FFiK)(2) showing superior self-assembling properties. The (FFiK)(2) hydrogels were used as scaffolds for cell culture, supporting cell proliferation without significant cytotoxicity, making them beneficial for cell engineering applications.
PEPTIDE SCIENCE (2021)
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Article Chemistry, Medicinal
Selection of fluorescent biosensors against galectin-3 from an NBD-modified phage library displaying designed α-helical peptides
Masahiro Hashimoto, Takayuki Miki, Iou Ven Chang, Hiroshi Tsutsumi, Hisakazu Mihara
Summary: This study demonstrated the selection of biosensors that selectively bind to Gal-3 from an NBD-modified peptide phage library. The fluorescence intensity of these biosensors increased with Gal-3 concentration, with a visible fluorescent response.
BIOORGANIC & MEDICINAL CHEMISTRY LETTERS (2021)
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Article Biochemistry & Molecular Biology
Biofunctional supramolecular hydrogels fabricated from a short self-assembling peptide modified with bioactive sequences for the 3D culture of breast cancer MCF-7 cells
Jyh Yea Chia, Takayuki Miki, Hisakazu Mihara, Hiroshi Tsutsumi
Summary: Short self-assembling peptide (FFiK)(2) and its derivatives can be functionalized by conjugating with various bioactive sequences to facilitate 3D culture of cancer cells. These functionalized peptide hydrogels can encapsulate MCF-7 cells without significant cytotoxicity, promoting their proliferation and potentially serving as a platform for drug screening targeting cancer stem cells.
BIOORGANIC & MEDICINAL CHEMISTRY (2021)
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Article Biochemical Research Methods
Effects of Hydrophobic Residues on the Intracellular Self-Assembly of De Novo Designed Peptide Tags and Their Orthogonality
Takayuki Miki, Keigo Kajiwara, Sae Nakayama, Masahiro Hashimoto, Hisakazu Mihara
Summary: Protein assemblies play vital roles in living systems, and self-assembling peptides (SAPs) serve as potential tags. In this study, we investigated the role of hydrophobic residues in SAPs and found that different hydrophobic amino acids significantly affect self-assembly propensity, assembly size, protein denaturation, and subcellular localization. Additionally, we explored the orthogonality of their interactions.
ACS SYNTHETIC BIOLOGY (2022)
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Article Biochemistry & Molecular Biology
Proximity labeling and identification of endogenous client proteins recruited to Y15-based artificial granules tethering a bait protein
Masahiro Hashimoto, Takayuki Miki, Tatsuya Niwa, Hisakazu Mihara
Summary: Protein clustering is a common phenomenon in cellular processes, where self-association of proteins regulates cellular signaling by recruiting downstream proteins. To understand these interactions, chemical biology tools are needed to identify proteins recruited to specific assemblies. In this study, we developed the IPRAG platform, which combines intracellular supramolecule construction with a proximity labeling method, to identify proteins recruited in artificial granules. Using Nck1 as a target protein, we successfully validated the IPRAG tool and identified proteins that interact with Nck1 directly or indirectly.
JOURNAL OF PEPTIDE SCIENCE (2023)
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Article Chemistry, Multidisciplinary
Pattern enrichment analysis for phage selection of stapled peptide ligands
Takayuki Miki, Keigo Namii, Kenta Seko, Shota Kakehi, Goshi Moro, Hisakazu Mihara
Summary: Phage display is a widely used technique for discovering peptides that bind to target proteins. This study introduces a new method called 'pattern enrichment analysis' to overcome challenges associated with compositional bias. By comparing sequence datasets from affinity-selected samples and non-selected samples, this method allows for the identification of enriched amino acid residue patterns and facilitates the discovery of peptide sequences and key binding residues. The effectiveness of this method was validated through screening against HDM2 protein and the SARS-CoV-2 spike RBD.
CHEMICAL SCIENCE (2022)
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Article Chemistry, Multidisciplinary
A guide-tag system controlling client enrichment into Y15 peptide-based granules for an in-cell protein recruitment assay
Takayuki Miki, Masahiro Hashimoto, Taichi Nakai, Hisakazu Mihara
Summary: The study developed a guide-tag system that concentrates client proteins into SAP-based scaffolds in cellular environments. This system provides a tool for analysing protein-protein interactions caused by protein clustering in cells.
CHEMICAL COMMUNICATIONS (2021)
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