Molecular simulations on electron transfer proteins

Cytochrome P450s are versatile biocatalysts with the ability to accept a vast range of substrates. However, to enable their high potential they are in need of electrons. Those are delivered via complex electron transfer chains including an electron donor (NAD(P)H) and one or two transfer proteins. Transfer proteins are exchangeable from di erent organisms and work in various combinations. To gain knowledge and understanding about possible interactions of redox partners and subsequent transport of electrons to cytochromes P450 (P450s) molecular modelling methods are vital. Increasing computational power makes in silico methods like molecular docking and molecular dynamics simulations more and more suitable to analyze biological systems like P450s. With these methods we were able to establish changes in progesterone hydroxylation and subsequently shift and increase the product spectrum. The introduction of modelled glycine and proline linker structures could help to explain the interactions of proteins and the accompanied electron transfer. Also we investigated combinations of redox partners to optimize the redox chain of CYP106A2. To lower the redox potential of adrenodoxin, a suitable redox partner of CYP106A2, methods of rational protein design were applied.Cytochrom P450 Enzyme sind breit einsetzbare Biokatalysatoren, die eine Vielzahl an Substraten akzeptieren k onnen, eine F ahigkeit die sie sehr attraktiv f ur eine biotechnologische Anwendung macht. Trotz ihrer vielseitigen Einsetzbarkeit gibt es jedoch einige Limitierungen, die es zu uberwinden gilt. Hierzu z ahlen beispielsweise die komplexen Elektronentransferprozesse oder die niedrige Elektronentransferrate. Die zur Elektronen ubertragung ben otigten Transferpartner sind uber verschiedene Organismen hinweg austauschbar und kombinierbar. Mit stetiger Erh ohung der Rechenleistungen werden in silico Methoden wie molekulares Docking oder Molek uldynamiksimulationen zur Analyse biologischer Systeme immer leichter einsetzbar. Im Rahmen unserer Forschungsarbeiten konnten wir damit Anderungen der Progesteronhydroxylierung bewirken, die das Produktspektrum verschob und den Ertrag wesentlich erh ohte. Die Modellierung von Glycin- und Prolin-Linker- Strukturen trug dazu bei, Interaktionen der Redoxpartner und den einhergehenden Elektronentransfer zu erkl aren. Des Weiteren wurden verschiedene Kombinationsm oglichkeiten von Redoxpartnern zur Optimierung der Redoxkette von CYP106A2 untersucht. Um das Redoxpotential von Adrenodoxin, einem passenden Redoxpartner des CYP106A2, herabzusetzen, kamen Methoden des rationalen Proteindesigns zum Einsatz

Effect of Oriented Electric Fields on Biologically Relevant Iron–Sulfur Clusters and Bioinformatics Investigations of Biotin Synthase

Enzymes, as biological catalysts, enjoy several benefits over the more commonly used metal catalysts in chemistry, particularly in terms of sustainability. They can, however, be more complicated to utilise and manipulate and research tends to focus on engineering enzymes for specific tasks where the complexity is reduced and a by-product of this is increased understanding of sequence-structure-function relationship. An alternative approach is to find broader problems whose solutions could be applied to the engineering of many enzymes, or at least a large, multipurpose superfamily of them. An excellent target for this type of approach is the radical S-adenosylmethionine (rSAM) superfamily, particularly due to its common mechanism of generating a radical species and using careful substrate control to dictate the reaction products across the different enzymes. This common mechanism includes an iron-sulfur cluster which can be influenced by the electrostatic environment, providing a clear path for study and promising powerful engineering opportunities. The focus of this research is an analysis of the effect of oriented electric fields on several relevant iron-sulfur clusters using a systematic, high throughput density-functional theory (DFT) study to gain both quantitative and qualitative information on the relative energies of spin states, orbitals, vertical electron affinities and spin-coupling constants. In addition, methods are identified for coupling this type of study with bioinformatic information for the purpose of enzyme engineering. Applying this to an exemplar of the rSAM superfamily, biotin synthase (BioB), indicates promising scope for variation at iron-sulfur cluster binding sites, whilst retaining functionality. Both the DFT results and the bioinformatics analysis represent a promising step towards the potential automation of enzyme engineering and is not limited to biotin synthase or even rSAM enzymes. This could result in improved development of a wide variety of chemical products in sustainable, efficient, and low-carbon syntheses, with the concomitant contributions to mitigating climate change

Effect of Oriented Electric Fields on Biologically Relevant Iron–Sulfur Clusters and Bioinformatics Investigations of Biotin Synthase

Enzymes, as biological catalysts, enjoy several benefits over the more commonly used metal catalysts in chemistry, particularly in terms of sustainability. They can, however, be more complicated to utilise and manipulate and research tends to focus on engineering enzymes for specific tasks where the complexity is reduced and a by-product of this is increased understanding of sequence-structure-function relationship. An alternative approach is to find broader problems whose solutions could be applied to the engineering of many enzymes, or at least a large, multipurpose superfamily of them. An excellent target for this type of approach is the radical S-adenosylmethionine (rSAM) superfamily, particularly due to its common mechanism of generating a radical species and using careful substrate control to dictate the reaction products across the different enzymes. This common mechanism includes an iron-sulfur cluster which can be influenced by the electrostatic environment, providing a clear path for study and promising powerful engineering opportunities. The focus of this research is an analysis of the effect of oriented electric fields on several relevant iron-sulfur clusters using a systematic, high throughput density-functional theory (DFT) study to gain both quantitative and qualitative information on the relative energies of spin states, orbitals, vertical electron affinities and spin-coupling constants. In addition, methods are identified for coupling this type of study with bioinformatic information for the purpose of enzyme engineering. Applying this to an exemplar of the rSAM superfamily, biotin synthase (BioB), indicates promising scope for variation at iron-sulfur cluster binding sites, whilst retaining functionality. Both the DFT results and the bioinformatics analysis represent a promising step towards the potential automation of enzyme engineering and is not limited to biotin synthase or even rSAM enzymes. This could result in improved development of a wide variety of chemical products in sustainable, efficient, and low-carbon syntheses, with the concomitant contributions to mitigating climate change