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A Flexible Platform for the Application of SAM-dependent enzymes

Periodic Reporting for period 2 - AppSAM (A Flexible Platform for the Application of SAM-dependent enzymes)

Reporting period: 2019-04-01 to 2020-09-30

S-adenosylmethionine (SAM)-dependent enzymes play an important role in all kingdoms of life. SAM is used as a methyl donor for the methylation of small molecules, proteins, and ribonucleic acids by conventional methyltransferases. It is futher the cofactor for radical SAM enzymes that can catalyse a multitude of complex reactions, including methylations at unactivated positions. In the wide field of epigenetics, methylations are also an important marker, an example are methylations of nucleobases that are introduced by conventional and radical SAM methyltransferases. The reactions catalysed by SAM-dependent enzymes are highly interesting for technical application, e.g. the synthesis of selectively methylated building blocks for pharmaceuticals. Often, the corresponding synthetically-chemical version of the reaction uses highly toxic and cancerogenic compounds and is rather unselective. In the AppSAM project, we are developing strategies to integrate SAM-dependent enzyms in multi-enzyme cascades to facilitate their handling and efficiency for chemical synthesis. These systems will be also useful for the mechanistic-functional investigation of SAM-dependent enzymes, e.g. from epigenetic pathways or natural product biosynthesis, using modified (co-)substrates that can be produced in situ.
So far, the cyclic SAM regeneration system has been optimised regarding reactions conditions and turnover numbers. In addition, initial steps were made towards the extension of the system for other SAM-dependent enzymes.
The cyclic SAM regeneration system can be used for convential methyltransferases and will be extended to be applicable for further SAM-dependent enzymes. In addition to these in vitro systems, in vivo systems using whole cells will be explored with a range of enzymes using the SAM cofactor. The goal for the end of the AppSAM project is the establisment of a sustainable, flexible platform to use SAM-dependent enzymes for chemical synthesis, which can be easily adapted to new systems.