Dokument: Computational studies of enzymatic reactions: Elucidating the Taxadiene Synthase mechanism and the CalB-catalyzed hydrolysis of propranolol esters

Titel:	Computational studies of enzymatic reactions: Elucidating the Taxadiene Synthase mechanism and the CalB-catalyzed hydrolysis of propranolol esters
URL für Lesezeichen:	https://docserv.uni-duesseldorf.de/servlets/DocumentServlet?id=56782
URN (NBN):	urn:nbn:de:hbz:061-20210728-095430-5
Kollektion:	Dissertationen
Sprache:	Englisch
Dokumententyp:	Wissenschaftliche Abschlussarbeiten » Dissertation
Medientyp:	Text
Autor:	van Rijn, Jeannette [Autor]
Dateien:	[Dateien anzeigen] Adobe PDF [Details] 17,30 MB in einer Datei [ZIP-Datei erzeugen] Dateien vom 05.07.2021 / geändert 05.07.2021
Beitragende:	Prof. Dr. Neese, Frank [Gutachter] Prof. Dr. Marian, Christel [Gutachter]
Dewey Dezimal-Klassifikation:	500 Naturwissenschaften und Mathematik » 540 Chemie
Beschreibung:	Taxol (paclitaxel, an anticancer drug) and Propranolol (a beta-adrenergic blocking agent) are two important drugs for which insights into the reaction mechanism of the synthesis could open up the possibility to improve the production process through rational design of enzymes, mutations or better substrates. For Taxol, the current commercial production processes heavily depend on the taxus plant and produce significant toxic waste streams, making them less environmentally sustainable and increasing the cost of taxol. The first committed step in the production of Taxol is the conversion of geranylgeranyl diphosphate (GGPP) to taxadiene (T), catalyzed by taxadiene synthase (TXS). For this step we performed molecular dynamics (MD) simulations to study the dynamic behavior of noncovalent enzyme carbocation complexes. Taxadiene and the observed four side products originate from the deprotonation of carbocation intermediates. The MD simulations of the TXS carbocation complexes provide insights into potential deprotonation mechanisms of such carbocations, showing water bridges which may allow the formation of side products via multiple proton transfer reactions. Combined quantum mechanics/molecular mechanics (QM/MM) calculations were used to investigate energy profiles for the conversion of GGPP to T, as well as to minor products for different configurations of relevant TXS carbocation complexes. The QM/MM calculations suggest a reaction pathway for the conversion of GGPP to T, which slightly differs from previous proposals regarding the number of reaction steps and the conformation of the carbocations. The QM/MM results also indicate that formation of minor products via water-assisted deprotonation of the carbocations is highly exothermic, by about -7 to -23 kcal/mol. Curiously, however, the computed barriers and reaction energies indicate that the formation of some of the minor products is more facile than the formation of T. Thus, the present calculations provide detailed insights into possible reaction pathways and into the origin of the promiscuity of TXS, but they do not reproduce the product distribution observed experimentally. Propranolol is commercially available as a racemic mixture, though the S-enantiomer is significantly more active as a drug than both the R-enantiomer and the racemic mixture, while the racemic mixture has been shown to cause severe side effects. A possible way to produce enantiomerically pure Propranolol is the use of lipase catalyzed hydrolysis reactions of ester compounds. The CalB-catalyzed hydrolysis of propranolol esters was investigated for a range of acyl donors of various structures and different lengths to understand how the structure of the acyl donor affects the binding of the propranolol ester with CalB, the reactivity and the enantioselectivity. Docking results suggests that acyl donors with branched alkyl chains are too sterically demanding to be reactive. Subsequent molecular simulations of the propranolol esters with linear chains suggest the reactivity of propranolol esters with shorter chains (O-acetyl-propranolol, M0) to be high compared to propranolol esters with longer ones (O-propanoyl-propranolol, M1 and O-butanoyl-propranolol, M2). %This based on the more favorable binding process for M0; the reduced lifetime of productive noncovalent enzyme-substrate complexes (Michaelis complex, MCC) for both the R- and S-enantiomer of M1 and M2 in the MD runs and the fact that for M1 and M2 reactive MCCs were only identified in one of the two binding modes. The MD simulations also suggest that the hydrolysis reaction of racemic (R,S)-O-acetyl-propranolol (M0) will prefer to generate R-propranolol. In agreement with MD results, the QM/MM calculations of the hydrolysis reaction of M0 suggests an enantiomeric preference for the R-product. The activation energy gap between the reaction of R- and S-M0 is 6.2 kcal/mol, which is larger than that of the acylation reaction, indicating a potentially higher enantioselectivity for the hydrolysis reaction.
Lizenz:	Urheberrechtsschutz
Fachbereich / Einrichtung:	Mathematisch- Naturwissenschaftliche Fakultät
Dokument erstellt am:	28.07.2021
Dateien geändert am:	28.07.2021
Promotionsantrag am:	11.09.2020
Datum der Promotion:	07.12.2020