Article
Biochemistry & Molecular Biology
Pete Heinzelman, Philip A. A. Romero
Summary: ACE2 variants with improved specificity towards Ang-II and wild-type or greater Ang-II hydrolytic activity were discovered using yeast display-based liquid chromatography screen and directed evolution, which have potential as therapeutic agents for multiple human diseases.
Article
Biochemical Research Methods
Elisabeth Laurent, Anna Sieber, Benjamin Salzer, Anna Wachernig, Jacqueline Seigner, Manfred Lehner, Rene Geyeregger, Bernhard Kratzer, Ulrich Jager, Renate Kunert, Winfried F. Pickl, Michael W. Traxlmayr
Summary: Engineered stable CD19-ECD variants (SuperFolder) have been developed, with improved expression rates and efficient purification in monomeric forms, overcoming obstacles in the development and analysis of CD19-targeted therapeutics. These mutants preserve the wild type sequence (>98.8%) and hold promise for a range of applications in basic immunology and CD19-targeted cancer immunotherapy.
ACS SYNTHETIC BIOLOGY
(2021)
Article
Immunology
Desislava Yanakieva, Lena Vollmer, Andreas Evers, Vanessa Siegmund, Paul Arras, Lukas Pekar, Achim Doerner, Bernhard Valldorf, Harald Kolmar, Stefan Zielonka, Simon Krah
Summary: In this study, a novel symmetric bispecific antibody format was presented by grafting cattle-derived knob paratopes onto peripheral loops of the IgG1 Fc region. Knob architectures obtained from bovine ultralong CDR-H3 antibodies were inserted into the AB loop or EF loop of the CH3 domain to introduce an artificial binding specificity into an IgG molecule. The generated bispecific antibodies demonstrated retained binding affinities of the inserted knob domains and versatile effector cell redirection functionalities without compromising Fc mediated functions such as FcRn or Fc gamma RIIIa binding.
FRONTIERS IN IMMUNOLOGY
(2023)
Article
Biochemical Research Methods
Wen-Ching Lin, Hao-Cheng Tang, Han Ying Wang, Chia-Yi Kao, You-Chiun Chang, Athena Hsu Li, Shi-Bing Yang, Kurt Yun Mou
Summary: This study introduces a unique mutagenesis method called the reverse Kunkel method, which combines the advantages of user-defined mutagenesis and random mutagenesis. By coupling with phage display and yeast display selections, the method successfully generates improved antibodies with enhanced affinity and immunostaining performance.
ACS SYNTHETIC BIOLOGY
(2022)
Review
Chemistry, Multidisciplinary
Yajie Wang, Pu Xue, Mingfeng Cao, Tianhao Yu, Stephan T. Lane, Huimin Zhao
Summary: Directed evolution aims to accelerate the natural evolution process of biological molecules and systems through gene diversification and library screening, serving as a powerful tool for engineering improved functions in proteins, metabolic pathways, and whole genomes. Common strategies include gene diversification, screening/selection methods, and continuous evolution, with applications in nucleic acids, proteins, pathways, genetic circuits, viruses, and cells. Challenges and future perspectives in directed evolution are also discussed.
Article
Chemistry, Multidisciplinary
Anna Camus, Gisele Truong, Peer R. . E. Mittl, Greta Markert, Donald Hilvert
Summary: This study demonstrates the successful switch of substrate specificity in gatekeeper adenylation domains of nonribosomal peptide synthetases using high-throughput engineering, leading to efficient production of diverse peptide metabolites.
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2022)
Article
Chemistry, Multidisciplinary
Carter J. Teal, Marian H. Hettiaratchi, Margaret T. Ho, Arturo Ortin-Martinez, Ahil N. Ganesh, Andrew J. Pickering, Alex W. Golinski, Benjamin J. Hackel, Valerie A. Wallace, Molly S. Shoichet
Summary: By using directed evolution, specific affibody binding partners have been engineered to independently control the release rates of therapeutic proteins. In vivo testing on mice demonstrated the sustained activity of the controlled release, leading to improved efficacy.
ADVANCED MATERIALS
(2022)
Article
Biochemistry & Molecular Biology
Kosuke Oyama, Takatoshi Ohkuri, Jinta Ochi, Jose M. M. Caaveiro, Tadashi Ueda
Summary: The glycosylation and stabilization of the CH2 domain in IgG can decrease its aggregation and immunogenicity, and the combination of these two modifications can completely suppress its aggregation behavior.
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
(2021)
Article
Biotechnology & Applied Microbiology
Wenjia Lou, Samuel D. Stimple, Alec A. Desai, Emily K. Makowski, Sibel Kalyoncu, Jesper E. Mogensen, Lotte T. Spang, Desiree J. Asgreen, Arne Staby, Karen Duus, Jan Amstrup, Yulei Zhang, Peter M. Tessier
Summary: Biologics such as peptides and proteins have attractive attributes, but are prone to aggregation. By developing conformation-specific antibodies, a sensitive immunoassay for detecting aggregates has been successfully created. These antibodies not only exhibit high conformational specificity, but also cross-react with amyloid fibrils formed by various polypeptides.
BIOTECHNOLOGY AND BIOENGINEERING
(2021)
Review
Biotechnology & Applied Microbiology
Zhanzhi Liu, Sheng Chen, Jing Wu
Summary: Inspired by natural evolution, directed evolution mutates genes randomly under artificial evolution conditions and screens variants for desired properties. Directed evolution is crucial for enhancing protein properties and involves the construction of diverse libraries and efficient screening methods. This review organizes ultrahigh-throughput screening principles based on protein properties and provides a comprehensive introduction to the latest progress and future trends in ultrahigh-throughput screening technologies for directed evolution.
TRENDS IN BIOTECHNOLOGY
(2023)
Review
Chemistry, Multidisciplinary
Romany J. McLure, Sheena E. Radford, David J. Brockwell
Summary: Directed evolution is a powerful tool for engineering new functions in biomolecules and gaining insights into protein behavior. It accelerates the process of natural evolution through multiple rounds of gene diversification and selection.
TRENDS IN CHEMISTRY
(2022)
Article
Chemistry, Medicinal
Muriel D. van Kampen, Leonie H. A. M. Kuipers-De Wilt, Marielle L. van Egmond, Petra Reinders-Blankert, Ewald T. J. van den Bremer, Guanbo Wang, Albert J. R. Heck, Paul W. H. Parren, Frank J. Beurskens, Janine Schuurman, Rob N. de Jong
Summary: This study biophysically characterized the impact of self-association in IgG antibody variants, finding a correlation between increased self-association propensity and functional activity. The mutation E430G decreased conformational stability, but had no substantial effect on storage stability. These findings provide insights for the design and development of IgG-based therapeutics.
JOURNAL OF PHARMACEUTICAL SCIENCES
(2022)
Article
Chemistry, Multidisciplinary
Jonathan Ruiz Esquius, David J. Morgan, Gerardo Algara Siller, Diego Gianolio, Matteo Aramini, Leopold Lahn, Olga Kasian, Simon A. Kondrat, Robert Schloegl, Graham J. Hutchings, Rosa Arrigo, Simon J. Freakley
Summary: The oxygen evolution reaction (OER) is crucial for future energy systems based on water electrolysis. Iridium oxides show promise as catalysts due to their corrosion resistance in acidic and oxidizing conditions. The transformation of highly active iridium (oxy)-hydroxides prepared using alkali metal bases into rutile IrO2 at high temperatures (>350 degrees C) results in poor activity, but the transformation into Li-intercalated IrOx retains comparative activity and improved stability. This nanocrystalline lithium iridate form could be more resistant to industrial procedures and help stabilize the high populations of redox active sites in amorphous iridium (oxy)hydroxides.
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2023)
Article
Biochemical Research Methods
Biaobiao Luo, Moonsoo M. Jin, Xiaohua Li, Nokwanda P. Makunga, Xuebo Hu
Summary: Enzymes immobilized through yeast surface display (YSD) offer a cost-effective method for enriching enzymes for biochemistry characterization and application. This study demonstrates the feasibility of reconstituting complex multi-enzymatic systems for chemical biosynthesis in vitro by exploiting YSD-immobilized cascade enzymes.
ACS SYNTHETIC BIOLOGY
(2021)
Review
Biochemistry & Molecular Biology
Alireza Shoari, Ghazaleh Khalili-Tanha, Mathew A. Coban, Evette S. Radisky
Summary: The study of protein-protein interactions (PPIs) and protein-based inhibitors often uses two strategies: combinatorial libraries and computational modeling. When combined, these strategies can generate superior outcomes in protein engineering, providing insight into PPI specificity and affinity.
FRONTIERS IN MOLECULAR BIOSCIENCES
(2023)
Article
Biochemistry & Molecular Biology
Kevin Nys, Paul Georg Furtmueller, Christian Obinger, Sabine Van Doorslaer, Vera Pfanzagl
Summary: This study investigates the formation of tyrosine radicals in B-type dye-decolorizing peroxidase (KpDyP) and identifies the unique interaction between Y247 and Y92 specific to enterobacterial species. This provides evidence for the existence of long-range electron transfer pathways in B-type DyPs.
Article
Biochemistry & Molecular Biology
David Hoffmann, Stefan Mereiter, Yoo Jin Oh, Vanessa Monteil, Elizabeth Elder, Rong Zhu, Daniel Canena, Lisa Hain, Elisabeth Laurent, Clemens Gruenwald-Gruber, Miriam Klausberger, Gustav Jonsson, Max J. Kellner, Maria Novatchkova, Melita Ticevic, Antoine Chabloz, Gerald Wirnsberger, Astrid Hagelkruys, Friedrich Altmann, Lukas Mach, Johannes Stadlmann, Chris Oostenbrink, Ali Mirazimi, Peter Hinterdorfer, Josef M. Penninger
Summary: The study found that Clec4g and CD209c lectins can bind to the Spike protein of SARS-CoV-2, interfering with the virus' attachment to cell surfaces and significantly reducing infections. This provides hope for pan-variant therapeutic interventions.
Article
Biochemistry & Molecular Biology
Kevin Nys, Vera Pfanzagl, Jeroen Roefs, Christian Obinger, Sabine Van Doorslaer
Summary: This study used spectroscopic techniques to investigate the effect of active site perturbations on the structure of DyP enzymes from Klebsiella pneumoniae, highlighting the importance of key amino acids in maintaining heme distal architecture. Additionally, the study showed the significant impact of buffer choice and addition of cryoprotectant on experimental results.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2021)
Article
Plant Sciences
Yun-Ji Shin, Julia Koenig-Beihammer, Ulrike Vavra, Jennifer Schwestka, Nikolaus F. Kienzl, Miriam Klausberger, Elisabeth Laurent, Clemens Gruenwald-Gruber, Klemens Vierlinger, Manuela Hofner, Emmanuel Margolin, Andreas Weinhaeusel, Eva Stoeger, Lukas Mach, Richard Strasser
Summary: Studies on expressing glycosylated receptor binding domain (RBD) of SARS-CoV-2 spike protein in Nicotiana benthamiana revealed that a truncated RBD-215 variant can be efficiently expressed with glycosylation playing an important role in its functionality.
FRONTIERS IN PLANT SCIENCE
(2021)
Article
Biochemical Research Methods
Miriam Klausberger, Nikolaus F. Kienzl, Gerhard Stadlmayr, Clemens Grunwald-Gruber, Elisabeth Laurent, Katharina Stadlbauer, Florian Stracke, Klemens Vierlinger, Manuela Hofner, Gabriele Manhart, Wilhelm Gerner, Florian Grebien, Andreas Weinhausel, Lukas Mach, Gordana Wozniak-Knopp
Summary: This study aimed to optimize the receptor binding domain (RBD) of the SARS-CoV-2 spike protein produced in HEK293-6E cells. By modifying the O-glycosylation site and removing the unpaired cysteine residue, the researchers achieved a more stable and monomeric form of RBD. These optimized RBD variants showed excellent performance in detecting SARS-CoV-2 antibodies in convalescent sera, indicating their potential for serological testing and vaccine development.
BIOTECHNOLOGY JOURNAL
(2022)
Article
Biochemistry & Molecular Biology
Stefan Hofbauer, Marcello Pignataro, Marco Borsari, Carlo Augusto Bortolotti, Giulia Di Rocco, Gianina Ravenscroft, Paul G. Furtmuller, Christian Obinger, Marco Sola, Gianantonio Battistuzzi
Summary: The autosomal dominant striated muscle disease myoglobinopathy is caused by a single point mutation in human myoglobin, leading to increased reactivity towards hydrogen peroxide, enhanced tendency to form high-molecular-weight aggregates, and accelerated heme bleaching. These effects contribute to impaired oxygen binding capacity and faster heme dissociation, ultimately impacting the pathological symptoms of myoglobinopathy.
Article
Biology
Tuemay Capraz, Nikolaus F. Kienzl, Elisabeth Laurent, Jan W. Perthold, Esther Foederl-Hoebenreich, Clemens Gruenwald-Gruber, Daniel Maresch, Vanessa Monteil, Janine Niederhofer, Gerald Wirnsberger, Ali Mirazimi, Kurt Zatloukal, Lukas Mach, Josef M. Penninger, Chris Oostenbrink, Johannes Stadlmann
Summary: The interaction between the Spike protein of SARS-CoV-2 and ACE2 on host cells is crucial for viral entry, with glycosylation playing a critical role. Engineering ACE2 glycosylation can enhance virus neutralization without affecting protein stability or activity. Removing all N-glycans from recombinant soluble human ACE2 can create a superior decoy receptor for effective COVID-19 treatment.
Editorial Material
Biochemistry & Molecular Biology
Christian Obinger
Summary: This paper presents the first purification and characterization of a manganese peroxidase, demonstrating its ability to oxidize various compounds using hydrogen peroxide and Mn(II). This discovery in 1985 laid the foundation for extensive research on the structure-function relationships, ecological roles, and industrial applications of manganese peroxidases.
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
(2022)
Article
Biochemistry & Molecular Biology
Katharina Stadlbauer, Peter Andorfer, Gerhard Stadlmayr, Florian Ruker, Gordana Wozniak-Knopp
Summary: Inhibition of complement activation by overexpressing complement-regulatory proteins can disguise cancer cells from the immune system, but may also reduce the efficacy of therapeutic antibodies. By selecting CD59-specific Fc fragments, a bispecific antibody was created that can induce higher specific cytotoxicity at a lower concentration.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2022)
Review
Biochemical Research Methods
Magdalena Teufl, Charlotte U. Zajc, Michael W. Traxlmayr
Summary: The stability of a protein is crucial for its applicability, but improving functionality often leads to destabilization. Inserting mutations for new or improved functions can result in destabilization, a common trade-off in protein evolution. Strategies like using stable parental proteins, minimizing destabilization during engineering, and repairing damaged mutants have been successful in overcoming this challenge in protein engineering.
ACS SYNTHETIC BIOLOGY
(2022)
Article
Biochemistry & Molecular Biology
Anja Wagner, Edgar Galicia-Andres, Magdalena Teufl, Lukas Gold, Christian Obinger, Peter Sykacek, Chris Oostenbrink, Michael W. Traxlmayr
Summary: By screening a mutated EGFR library, previously unknown activating mutations in EGFR were discovered. These mutations are not only located in the kinase domain of EGFR, but also in other regions, including the extracellular and transmembrane domains.
Article
Biochemistry & Molecular Biology
Daniel Schmidt, Nikolaus Falb, Ilenia Serra, Marzia Bellei, Vera Pfanzagl, Stefan Hofbauer, Sabine Van Doorslaer, Gianantonio Battistuzzi, Paul G. Furtmuller, Christian Obinger
Article
Biochemistry & Molecular Biology
Daniel R. Ramos, Paul G. Furtmueller, Christian Obinger, Angeles Pena-Gallego, Ignacio Perez-Juste, J. Arturo Santaballa
Summary: Electronic structure calculations using DFT were conducted to examine the influence of water molecules and protonation on the heme group of peroxidases in different redox and spin states. The study discusses shared geometries, spectroscopic properties, and thermodynamics of peroxidases. Computed Gibbs free energies suggest that the corresponding aquo complexes are not thermodynamically stable, supporting the five-coordinate Fe(III) center in native ferric peroxidases with a non-bonding water molecule. Protonation of the ferryl oxygen of compound II is found to be necessary and computed Gibbs free energies reveal pK(a) values of approximately 8.5-9.0 for compound II.
Article
Biochemistry & Molecular Biology
Aleksandra Rodak, Katharina Stadlbauer, Madhusudhan Reddy Bobbili, Oskar Smrzka, Florian Rueker, Gordana Wozniak Knopp
Summary: This study developed an antibody-drug conjugate that showed cytotoxicity for IgE-positive cells. The antibody had fast internalization and strict target specificity, providing a valuable starting point for the development of an anti-IgE cell-depleting agent.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2023)