Geschmack ist vorhersagbar: Mit FlavorMiner. FlavorMiner heißt das Tool, das IPB-Chemiker und Partner aus Kolumbien jüngst entwickelt haben. Das Programm kann, basierend auf maschinellem Lernen (KI), anhand der…
Seit Februar 2021 bietet Wolfgang Brandt, ehemaliger Leiter der Arbeitsgruppe Computerchemie am IPB, sein Citizen Science-Projekt zur Pilzbestimmung an. Dafür hat er in regelmäßigen Abständen öffentliche Vorträge zur Vielfalt…
Müllers, Y.; Sadr, A. S.; Schenderlein, M.; Pallab, N.; D. Davari, M.; Glebe, U.; Reifarth, M.;Acrylate‐derived RAFT polymers for enzyme hyperactivation – boosting the α‐chymotrypsin enzyme activity using tailor‐made poly(2‐carboxyethyl)acrylate (PCEA)ChemCatChem16e202301685(2024)DOI: 10.1002/cctc.202301685
We study the hyperactivation of α‐chymotrypsin (α‐ChT) using the acrylate polymer poly(2‐carboxyethyl) acrylate (PCEA) in comparison to the commonly used poly(acrylic acid) (PAA). The polymers are added during the enzymatic cleavage reaction of the substrate N‐glutaryl‐L‐phenylalanine p‐nitroanilide (GPNA). Enzyme activity assays reveal a pronounced enzyme hyperactivation capacity of PCEA, which reaches up to 950% activity enhancement, and is significantly superior to PAA (revealing an activity enhancement of approx. 450%). In a combined experimental and computational study, we investigate α‐ChT/polymer interactions to elucidate the hyperactivation mechanism of the enzyme. Isothermal titration calorimetry reveals a pronounced complexation between the polymer and the enzyme. Docking simulations reveal that binding of polymers significantly improves the binding affinity of GPNA to α‐ChT. Notably, a higher binding affinity is found for the α‐ChT/PCEA compared to the α‐ChT/PAA complex. Further molecular dynamics (MD) simulations reveal changes in the size of the active site in the enzyme/polymer complexes, with PCEA inducing a more pronounced alteration compared to PAA, facilitating an easier access for the substrate to the active site of α‐ChT.
Publikation
Ninck, S.; Halder, V.; Krahn, J. H.; Beisser, D.; Resch, S.; Dodds, I.; Scholtysik, R.; Bormann, J.; Sewald, L.; Gupta, M. D.; Heilmann, G.; Bhandari, D. D.; Morimoto, K.; Buscaill, P.; Hause, B.; van der Hoorn, R. A. L.; Kaschani, F.; Kaiser, M.;Chemoproteomics Reveals the Pan-HER Kinase Inhibitor Neratinib To Target an Arabidopsis Epoxide Hydrolase Related to Phytohormone SignalingACS Chem. Biol.181076-1088(2023)DOI: 10.1021/acschembio.2c00322
Plant phytohormone pathways are regulated by an intricate network of signaling components and modulators, many of which still remain unknown. Here, we report a forward chemical genetics approach for the identification of functional SA agonists in Arabidopsis thaliana that revealed Neratinib (Ner), a covalent pan-HER kinase inhibitor drug in humans, as a modulator of SA signaling. Instead of a protein kinase, chemoproteomics unveiled that Ner covalently modifies a surface-exposed cysteine residue of Arabidopsis epoxide hydrolase isoform 7 (AtEH7), thereby triggering its allosteric inhibition. Physiologically, the Ner application induces jasmonate metabolism in an AtEH7-dependent manner as an early response. In addition, it modulates PATHOGENESIS RELATED 1 (PR1) expression as a hallmark of SA signaling activation as a later effect. AtEH7, however, is not the exclusive target for this physiological readout induced by Ner. Although the underlying molecular mechanisms of AtEH7-dependent modulation of jasmonate signaling and Ner-induced PR1-dependent activation of SA signaling and thus defense response regulation remain unknown, our present work illustrates the powerful combination of forward chemical genetics and chemical proteomics for identifying novel phytohormone signaling modulatory factors. It also suggests that marginally explored metabolic enzymes such as epoxide hydrolases may have further physiological roles in modulating signaling.
Publikation
Rajakumara, E.; Abhishek, S.; Nitin, K.; Saniya, D.; Bajaj, P.; Schwaneberg, U.; Davari, M. D.;Structure and cooperativity in substrate–enzyme interactions: Perspectives on enzyme engineering and inhibitor designACS Chem. Biol.17266-280(2022)DOI: 10.1021/acschembio.1c00500
Enzyme-based synthetic chemistry provides a green way to synthesize industrially important chemical scaffolds and provides incomparable substrate specificity and unmatched stereo-, regio-, and chemoselective product formation. However, using biocatalysts at an industrial scale has its challenges, like their narrow substrate scope, limited stability in large-scale one-pot reactions, and low expression levels. These limitations can be overcome by engineering and fine-tuning these biocatalysts using advanced protein engineering methods. A detailed understanding of the enzyme structure and catalytic mechanism and its structure–function relationship, cooperativity in binding of substrates, and dynamics of substrate–enzyme–cofactor complexes is essential for rational enzyme engineering for a specific purpose. This Review covers all these aspects along with an in-depth categorization of various industrially and pharmaceutically crucial bisubstrate enzymes based on their reaction mechanisms and their active site and substrate/cofactor-binding site structures. As the bisubstrate enzymes constitute around 60% of the known industrially important enzymes, studying their mechanism of actions and structure–activity relationship gives significant insight into deciding the targets for protein engineering for developing industrial biocatalysts. Thus, this Review is focused on providing a comprehensive knowledge of the bisubstrate enzymes’ structure, their mechanisms, and protein engineering approaches to develop them into industrial biocatalysts.
Publikation
Dippe, M.; Davari, M. D.; Weigel, B.; Heinke, R.; Vogt, T.; Wessjohann, L. A.;Altering the regiospecificity of a catechol
O‐methyltransferase through rational design: Vanilloid vs. isovanilloid motifs in the B‐ring of flavonoidsChemCatChem14e202200511(2022)DOI: 10.1002/cctc.202200511
Rational re-design of the substrate pocket of phenylpropanoid-flavonoid O-methyltransferase (PFOMT) from Mesembryanthe-mum crystallinum, an enzyme that selectively methylates the 3’-position (= meta-position) in catechol-moieties of flavonoids to guiacol-moieties, provided the basis for the generation of variants with opposite, i. e. 4’- (para-) regioselectivity and enhanced catalytic efficiency. A double variant (Y51R/N202W) identified through a newly developed colorimetric assay efficiently modified the para-position in flavanone and flavano-nol substrates, providing access to the sweetener molecule hesperetin and other rare plant flavonoids having an isovanil-loid motif.