Figure 3 - Biocatalytic cascade reaction including UPOs to retain an API Note : Adapted from Heckmann et al .
revolutionise the chemical industry . 1 In this work , Aminoverse ’ s UPO kit was used to identify a suitable biocatalyst for a three-step biocatalytic reaction to synthesise the chiral compound and the API ( 1S , 3R ) -3-hydroxycycloh exanecarbonitrile .
Starting from cyclohexene-1-nitril , the first step is the overoxidisation of the non-activated C-H in cyclohexene-1-nitril towards the ketone 3-oxocyclohexane-1- carbonitrile . This overoxidation is achieved best by UPO-O , with over 80 % conversion . The intermediate is then converted to the final API by an ene reductase and an alcohol dehydrogenase ( Figure 3 ).
The authors also investigated the stereoselectivity of the tested UPO , emphasising its diversity . In the first reaction using cyclohexene-1-nitrile as substrate , six R-selective and two S-selective UPOs were found to yield 30 – 70 % of the corresponding 3-hydroxycyclohex-1-ene-1- carbonitrile enantiomer under nonoptimised conditions .
Notably , the UPO with the highest overoxidation activity ( UPO-O ) is not the same as the one with the highest hydroxylation activity ( UPO-K for R , UPO-C for S ). Depending on the desired oxyfunctionalisation and substrate , a different UPO may turn out to be the most suitable biocatalyst , which illustrates the benefit of a panel screening .
KGO-catalysed reaction
The launch of the world ’ s first KGO kit by Aminoverse provides a means of synthesising new molecules that were even inaccessible with UPOs . A study by Hara et al . showed how KGOs can impact the search for a range of APIs . 2
As industry often uses ( 2S , 4R ) - 4-hydroxypyrrolidine-2-carboxylic acid ( L-hydroxyproline ), its isomer , ( 2S , 3S ) -4-hydroxypyrrolidine- 2-carboxylic ( trans-3-Lhydroxyproline ), represents an important building block for further synthesis . By using various biocatalysts , it was possible to produce the three other isomers : ( 2S , 4R ) -4-hydroxypyrrolidine- 2-carboxylic acid , ( 2S , 4S ) -4- hydroxypyrrolidine-2-carboxylic acid and ( 2S , 3R ) -4-hydroxypyrrolidine-2- carboxylic .
The lack of a highly selective enzyme made the biocatalytic production of trans-3-Lhydroxyproline inconceivable .
However , Hara et al . discovered one KGO biocatalyst that could synthesise trans-3-L-hydroxyproline from L-proline with 90 % conversion under the given reaction conditions ( Figure 4 ).
This study shows that nature ' s toolbox contains biocatalysts of incredible value for industrial applications . Biocatalyst kits like this can thus open access to cutting-edge synthesis routes , new intermediates , and novel products by significantly accelerating the timeline towards identification of an optimal catalyst
Limits & potential of biocatalysis
While a steadily growing number of companies start and expand the use of biocatalysis , the worldwide adoption across all chemical industries is still facing obstacles . The main concerns for PIs and research managers are costs , IP concerns , short-term availability and development times .
Biocatalysts are typically produced in biological cells , requiring some isolation of the catalyst prior to its use in a chemical process . These two steps of production and isolation account for the largest cost factor and are typically offset by increasing production scale .
60 SPECIALITY CHEMICALS MAGAZINE ESTABLISHED 1981