The opium poppy, Papaver somniferum, has been a source of medicinal alkaloids since the earliest civilizations; ca. 3400 B.C. The benzylisoquinoline alkaloid noscapine is produced commercially in P. somniferum for use as a cough suppressant and it also has potential as an anticancer compound. The first committed step in the recently elucidated noscapine biosynthetic pathway involves the conversion of scoulerine to tetrahydrocolumbamine by 9-O-methylation, catalysed by O-methyltransferase 1 (PSMT1). We demonstrate, through protein structures (obtained through rational crystal engineering at resolutions from 1.5 to 1.2Å for the engineered variants) across the reaction coordinate, how domain closure allows specific methyl transfer to generate the product. SAM-dependent methyl transfer is central to myriad natural products in plants, analysis of amino-acid sequence, now taking the threedimensional structure of PSMT1 and low identity homologs into account, begins to shed light on the structural features that govern substrate specificity in these key, ubiquitous, plant enzymes. We propose how thus allowing prediction across the wide genomic resource.

Small molecules inducing protein degradation are important pharmacological tools to interrogate complex biology and are rapidly translating into clinical agents. However, to fully realise the potential of these molecules, selectivity remains a limiting challenge. Herein, we addressed the issue of selectivity in the design of CRL4CRBN recruiting PROteolysis TArgeting Chimeras (PROTACs). Thalidomide derivatives used to generate CRL4CRBN recruiting PROTACs have well described intrinsic monovalent degradation profiles by inducing the recruitment of neo‐substrates such as GSPT1, Ikaros and Aiolos. We leveraged structural insights from known CRL4CRBN neo‐substrates to attenuate and indeed remove this monovalent degradation function in well‐known CRL4CRBN molecular glues degraders, namely CC‐885 and Pomalidomide. We then applied these design principles on a previously published BRD9 PROTAC (dBRD9‐A) and generated an analogue with improved selectivity profile. Finally, we implemented a computational modelling pipeline to show that our degron blocking design does not impact PROTAC induced ternary complex formation. We believe that the tools and principles presented in this work will be valuable to support the development of targeted protein degradation.

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