Development and optimization of cascades to promote the application of enzymes as environment-friendly catalysts

This work is dedicated to a particular kind of enzyme cascade, namely one that shall promote the industrial application of S-adenosyl-L-methionine-dependent methyltransferases (MT) as a greener alternative to traditional chemical methylation strategies. In principle, MTs can act as individual catalysts, when provided with a suitable substrate and methyl source. However, the reaction equilibrium or a state of product inhibition may be reached relatively early. Moreover, the expensiveness and instability of the methyl donor S-adenosyl-L-methionine (SAM), make it inevitable to couple the reaction to a recycling system in order to reach an economic viability. The herein contrived SAM regeneration cascade is bio-inspired and relies on four different enzymes (in addition to the MT): the demethylated donor molecule shall be broken down to L-homocysteine and adenosine by an S-adenosyl-L-homocysteine hydrolase (SAHH). While the former will accumulate in the current setup, the latter is entering the recycling via a phosphorylation to adenosine monophosphate (AMP), catalysed by an adenosine kinase (AK). This is followed by further phosphorylations to adenosine triphosphate (ATP) through the activity of a polyphosphate kinase (PPK) and driven by an excess of inorganic polyphosphate (polyP). The last recycling step is the fusion of ATP and L-methionine in the catalytic site of a methionine adenosyltransferase (MAT), which yields SAM again.