Cytochrome P450 monooxygenases (P450s or CYPs) are a unique family of enzymes which are capable of catalysing the regio- and stereospecific oxidation of non-functionalised hydrocarbons. Despite the enormous synthetic potential of P450s, these enzymes have yet to be extensively employed for research purposes or in industry. Lack of stability, low activity, narrow substrate specificity, expensive cofactor requirements, limited solvent tolerance and electron supply are some of the main reasons why the academic and industrial implementation of these important biocatalysts remains a challenge. Considering the significance of P450s, many research groups have focused on improving their properties in an effort to make more robust catalysts with broad synthetic applications. This article focuses on some of the factors that have limited the exploitation of P450s and explores some of the significant steps that have been taken towards addressing these limitations. © 2011 The Royal Society of Chemistry.