Cytochrome P450s are a superfamily of haem-dependent enzymes that catalyse the oxidation of natural organic compounds, most commonly via insertion of an oxygen atom from atmospheric dioxygen into a carbon-hydrogen bond. Since there is widespread interest in the selective functionalisation of C-H bonds, P450s have been extensively studied and evolved, particularly P450BM3 from Bacillus megaterium, which is catalytically self-sufficient. In this thesis, a P450BM3 variant library has been developed based on four 'generic accelerators', which have showed enhanced activity towards a broad range of non-natural substrates such as polyaromatic hydrocarbons and terpenes. The library was refined to 48 variants based on the screening results in order to maintain diversity of substrate pocket topology for oxidising a broad coverage of organic molecules. Successful applications include drug metabolism, drug fragment diversification, synthesis of flavour and fragrance compounds, steroid oxidation, late-stage functionalisation in total synthesis, and C-H amination. Total conversion was observed in most cases with a sufficient turnover number that enabled all the products to be fully characterised by NMR and MS. Two novel P450 activities were discovered as oxidative decarboxylation of an a-hydroxy carboxylic acid and C-H amination. The substrate range and varied product profiles suggest that this enzyme library is a good basis for developing selective C-H activation catalysts while expanding the P450 catalytic repertoire to include reactions important in synthetic chemistry, potentially opening new opportunities for biocatalysis.