Evolved enzymes for efficient chemoenzymatic routes in the synthesis of APIs
Dr Osama Mahmoud and Dr Thomas Daussmann of Enzymaster present recent examples of biocatalyst development for sustainable and stereoselective synthesis of API intermediates of drugs against neurological diseases and disorders *
Neurological diseases and disorders are a major and increasing global health challenge , which account for a substantial portion of the disease burden worldwide . 1 In recent years , several drugs have been developed to restore , minimise or compensate the functional deficits of patients , aiming at improving the quality of life .
Recent advances in enzyme discovery and protein engineering have paved the way for the development of new , efficient routes for drug synthesis outcompeting purely chemical approaches . Additionally , syntheses including enzymatic steps are in most cases more sustainable and environmentally friendly . 2
Figure 1 - Enzymatic synthesis of Ubrogepant IM
Recent outstanding examples are the production of tert-butyl (( 5S , 6R ) -6- methyl-2-oxo-5-phenylpiperidin-3-yl ) carbamate ( Ubrogepant intermediate ) and ( S ) -3-isobutyl-γ-aminobutyric acid ( Pregabalin ). In both cases directed evolution was performed using Enzymaster ’ s proprietary BioEngine ** platform guided by our BioNavigator ** bioinformatics toolbox for smart enzyme library design .
BioEngine is a directed evolution platform with integrated computational enzyme engineering . Based on the computational results , smart and small libraries of enzyme variants are constructed , which can be conveniently screened via HPLC , GC or MS to characterise a comprehensive range of desired properties for these variants under process conditions .
This makes it possible to reach biocatalytic process targets in typically only three to six rounds of combined in silico and experimental directed evolution , which corresponds to a timeline of four to eight months .
The BioNavigator toolbox is a set of advanced methods for computational enzyme identification , in silico activity and selectivity prediction , and stability calculation under process conditions . This includes bioinformatics-driven hot spot identification , focused combinatorial library design and in silico loop design .
AI and structure‐based recombination algorithms are also applied for in silico pre-screening to enhance the combinatorial sequence diversity coverage beyond traditional random and sitesaturation mutagenesis approaches , thereby increasing the sequence space coverage ( up to 106 variants / day for activity , selectivity and stability ) and identifying all important sequence areas .
Ubrogepant intermediate synthesis via evolved transaminase
Ubrogepant is a CGRP receptor antagonist API that has been approved for the treatment of migraine by the FDA in 2019 . The major challenge in the manufacture of this compound is the preparation of the key coupling partners – an amino lactam and a spiro
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