BIOMAT Database Logo BIOMAT Database Logo

A single active site mutation inverts stereoselectivity of 16-hydroxylation of testosterone catalyzed by engineered cytochrome P450 BM3. 2012

Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Department of Chemistry and Pharmaceutical Sciences, Division of Molecular and Computational Toxicology, Vrije Universiteit, De Boelelaan 1083, 1081 HV Amsterdam, Netherlands.

Inversion of stereoselectivity: screening of a minimal mutant library revealed a cytochrome P450 BM3 variant M01 A82W S72I capable of producing 16 α-OH-testosterone. Remarkably, a single active site mutation S72I in M01 A82W inverted the stereoselectivity of hydroxylation from 16 β to 16 α. Introduction of S72I mutation in another 16 β-OH-selective variant M11 V87I, also resulted in similar inversion of stereoselectivity.

UI MeSH Term Description Entries
D009154 Mutation Any detectable and heritable change in the genetic material that causes a change in the GENOTYPE and which is transmitted to daughter cells and to succeeding generations. Mutations
D003577 Cytochrome P-450 Enzyme System A superfamily of hundreds of closely related HEMEPROTEINS found throughout the phylogenetic spectrum, from animals, plants, fungi, to bacteria. They include numerous complex monooxygenases (MIXED FUNCTION OXYGENASES). In animals, these P-450 enzymes serve two major functions: (1) biosynthesis of steroids, fatty acids, and bile acids; (2) metabolism of endogenous and a wide variety of exogenous substrates, such as toxins and drugs (BIOTRANSFORMATION). They are classified, according to their sequence similarities rather than functions, into CYP gene families (>40% homology) and subfamilies (>59% homology). For example, enzymes from the CYP1, CYP2, and CYP3 gene families are responsible for most drug metabolism. Cytochrome P-450,Cytochrome P-450 Enzyme,Cytochrome P-450-Dependent Monooxygenase,P-450 Enzyme,P450 Enzyme,CYP450 Family,CYP450 Superfamily,Cytochrome P-450 Enzymes,Cytochrome P-450 Families,Cytochrome P-450 Monooxygenase,Cytochrome P-450 Oxygenase,Cytochrome P-450 Superfamily,Cytochrome P450,Cytochrome P450 Superfamily,Cytochrome p450 Families,P-450 Enzymes,P450 Enzymes,Cytochrome P 450,Cytochrome P 450 Dependent Monooxygenase,Cytochrome P 450 Enzyme,Cytochrome P 450 Enzyme System,Cytochrome P 450 Enzymes,Cytochrome P 450 Families,Cytochrome P 450 Monooxygenase,Cytochrome P 450 Oxygenase,Cytochrome P 450 Superfamily,Enzyme, Cytochrome P-450,Enzyme, P-450,Enzyme, P450,Enzymes, Cytochrome P-450,Enzymes, P-450,Enzymes, P450,Monooxygenase, Cytochrome P-450,Monooxygenase, Cytochrome P-450-Dependent,P 450 Enzyme,P 450 Enzymes,P-450 Enzyme, Cytochrome,P-450 Enzymes, Cytochrome,Superfamily, CYP450,Superfamily, Cytochrome P-450,Superfamily, Cytochrome P450
D006900 Hydroxylation Placing of a hydroxyl group on a compound in a position where one did not exist before. (Stedman, 26th ed) Hydroxylations
D013237 Stereoisomerism The phenomenon whereby compounds whose molecules have the same number and kind of atoms and the same atomic arrangement, but differ in their spatial relationships. (From McGraw-Hill Dictionary of Scientific and Technical Terms, 5th ed) Molecular Stereochemistry,Stereoisomers,Stereochemistry, Molecular,Stereoisomer
D013739 Testosterone A potent androgenic steroid and major product secreted by the LEYDIG CELLS of the TESTIS. Its production is stimulated by LUTEINIZING HORMONE from the PITUITARY GLAND. In turn, testosterone exerts feedback control of the pituitary LH and FSH secretion. Depending on the tissues, testosterone can be further converted to DIHYDROTESTOSTERONE or ESTRADIOL. 17-beta-Hydroxy-4-Androsten-3-one,17-beta-Hydroxy-8 alpha-4-Androsten-3-one,8-Isotestosterone,AndroGel,Androderm,Andropatch,Androtop,Histerone,Sterotate,Sustanon,Testim,Testoderm,Testolin,Testopel,Testosterone Sulfate,17 beta Hydroxy 4 Androsten 3 one,17 beta Hydroxy 8 alpha 4 Androsten 3 one,8 Isotestosterone
D015202 Protein Engineering Procedures by which protein structure and function are changed or created in vitro by altering existing or synthesizing new structural genes that direct the synthesis of proteins with sought-after properties. Such procedures may include the design of MOLECULAR MODELS of proteins using COMPUTER GRAPHICS or other molecular modeling techniques; site-specific mutagenesis (MUTAGENESIS, SITE-SPECIFIC) of existing genes; and DIRECTED MOLECULAR EVOLUTION techniques to create new genes. Genetic Engineering of Proteins,Genetic Engineering, Protein,Proteins, Genetic Engineering,Engineering, Protein,Engineering, Protein Genetic,Protein Genetic Engineering
D055162 Biocatalysis The facilitation of biochemical reactions with the aid of naturally occurring catalysts such as ENZYMES.
D020134 Catalytic Domain The region of an enzyme that interacts with its substrate to cause the enzymatic reaction. Active Site,Catalytic Core,Catalytic Region,Catalytic Site,Catalytic Subunit,Reactive Site,Active Sites,Catalytic Cores,Catalytic Domains,Catalytic Regions,Catalytic Sites,Catalytic Subunits,Core, Catalytic,Cores, Catalytic,Domain, Catalytic,Domains, Catalytic,Reactive Sites,Region, Catalytic,Regions, Catalytic,Site, Active,Site, Catalytic,Site, Reactive,Sites, Active,Sites, Catalytic,Sites, Reactive,Subunit, Catalytic,Subunits, Catalytic

Related Publications

Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Hydroxylation of non-substituted polycyclic aromatic hydrocarbons by cytochrome P450 BM3 engineered by directed evolution.
March 2013, Journal of inorganic biochemistry,
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
P450-BM3-Catalyzed Sulfoxidation versus Hydroxylation: A Common or Two Different Catalytically Active Species?
January 2020, Journal of the American Chemical Society,
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
A single mutation in cytochrome P450 BM3 changes substrate orientation in a catalytic intermediate and the regiospecificity of hydroxylation.
February 1997, Biochemistry,
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Direct conversion of ethane to ethanol by engineered cytochrome P450 BM3.
October 2005, Chembiochem : a European journal of chemical biology,
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Active site substitution A82W improves the regioselectivity of steroid hydroxylation by cytochrome P450 BM3 mutants as rationalized by spin relaxation nuclear magnetic resonance studies.
January 2012, Biochemistry,
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Testosterone 1 beta-hydroxylation by human cytochrome P450 3A4.
October 2004, European journal of biochemistry,
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Nonsubstrate recognition site residues are involved in testosterone hydroxylation by cytochrome P450 CYP 2C11.
January 1999, Archives of biochemistry and biophysics,
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Alkane metabolism by cytochrome P450 BM3.
November 1993, Biochemical Society transactions,
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Engineered alkane-hydroxylating cytochrome P450(BM3) exhibiting nativelike catalytic properties.
January 2007, Angewandte Chemie (International ed. in English),
Harini Venkataraman, and Stephanie B A de Beer, and Laura A H van Bergen, and Nick van Essen, and Daan P Geerke, and Nico P E Vermeulen, and Jan N M Commandeur
Regio- and stereo-selective 1β-hydroxylation of lithocholic acid by cytochrome P450 BM3 mutants.
August 2023, Biotechnology and bioengineering,
Copied contents to your clipboard!