PCR has transformed the field of biotechnology due to its ability to amplify small concentrations of nucleic acids in an autocatalytic manner.   A similar method to amplify enzymes in an autocatalytic �PCR� like process has the potential to revolutionize biotechnology, due to the large number of assays and procedures that use enzyme activity, ranging from ELISAs to targeted drug delivery.  In this report we present an enzyme amplification strategy, termed SAD (Self Accelerating Dimerization), which can amplify beta lactamase enzymatic activity by 9 orders of magnitude, via an autocatalytic process that uses beta lactamase activity to generate more beta lactamase.   SAD is based on an auto-catalytic loop generated from split proteins that dimerize in the presence of rapamycin, and is engineered so that rapamycin generates beta lactamase and beta lactamase then produces more rapamycin.  We show here that SAD causes exponential growth in beta-lactamase activity, and can amplify femtomolar concentrations of beta lactamase to micromolar levels within minutes.  The SAD process has numerous applications, and we demonstrate here that SAD can enable the visual detection of  5CFUs of beta lactamase expressing bacteria, via the amplification of beta lactamase activity.  The SAD process has the potential to have a significant impact on the biomedical sciences, given the widespread use of beta lactamase.

Autocatalytic amplification of enzyme activity using split proteins

247th National Meeting (2014)

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