# Recent Advances in Biocatalysis for Drug Synthesis
## 2 Biocatalysis for Drug Synthesis
### Expansion of Product Spectrum
#### 2.1.1 Natural Metabolism as Source of Enzymes and Biochemical Pathways
\begin{table}
\begin{tabular}{c c c c c c c c} \hline \hline Enzyme & \begin{tabular}{c} Source \\ Organism \\ \end{tabular} & Modification & Biocatalyst & Substrate & Product & \begin{tabular}{c} Process \\ Performance \\ \end{tabular} & Reference \\ \hline \begin{tabular}{c} Proline \\ hydroxylase \\ SmP4H \\ \end{tabular} & \begin{tabular}{c} _Sinorhizobium_ \\ _melidi_ \\ \end{tabular} & \begin{tabular}{c} One amino \\ acid exchange \\ (W40Y) \\ \end{tabular} & \begin{tabular}{c} Isolated \\ enzyme \\ \end{tabular} & \begin{tabular}{c} L-Homophen- \\ polaramine \\ \end{tabular} & \begin{tabular}{c} 3,4-Desaturated \\ l-homophen- \\ ylalanine \\ \end{tabular} & \begin{tabular}{c} \(\mathrm{k_{cat}}\) \\ \(0.83\pm 0.02\) min\({}^{-1}\), \\ 50.7\% yield \\ \end{tabular} & [21] \\ \hline \begin{tabular}{c} Proline \\ hydroxylase \\ SmP4H \\ \end{tabular} & \
\begin{tabular}{c} _Sinorhizobium_ \\ _melidi_ \\ \end{tabular} &
\begin{tabular}{c} Engineered \\ variant (six \\ amino acid \\ exchanges) \\ \end{tabular} & Cell lysate & L-Proline & _cis_ & \\ \hline \hline \end{tabular}
\end{table} 
Table 1: _Cont._[PERSON] et al. found a way for the biocatalytic selective monoacylation of symmetrical diamines [18]. This is chemically challenging, as one amino group must be selectively protected/deprotected during the course of the synthesis. [PERSON] et al. detected the ability of carboxylic acid reductases (CARs) to catalyze amide bond formations [30]. [PERSON] et al. employed a truncated version of a carboxylic acid reductase from _Mycobacterium marinum_ denoted CARmm-A [18]. The reaction was optimized by increasing the substrate loading (1, 5, and 10 mM) and the formerly-needed excess of the amine could be lowered from 100 to 17.5 equivalents. To overcome the issue of stoichiometrically-needed ATP, a regeneration system, in the form of a polyphosphate kinase class III from _Cytoplasna lutchisonii_, was successfully employed. With the depicted system in hand, a whole range of products was accessible in a single biocatalytic step, including the APIs lazabemide (anti-Parkinson) and cinepazide (vasodilator) (Figure 2C). Figure 2: Novel biocatalytic routes to access amides. (**A**) Synthesis of a key intermediate of losmapimod from 6-chloronicotinic acid, and neopentylamine utilizing a 4-chlorobenzene CL from _Alcaligenes_ sp. (CBL), and serotonin hydroxycinnamoyl transferase from _Capsicum annuum_ (66 CaT) [16]. (**B**) McbA catalyzed synthesis of moclobemide from _para_-chlorobenzoic acid, and 4-(2-aminoethyl)morpholine [17]. (**C**) Synthesis of cinepazide by the truncated carboxylic acid reductase from _Mycobacterium marinum_ CARmm-A [18]. Imine reductases (IREDs) are a relatively young class of enzymes, having been discovered around the year 2011 [31]. They are NADPH-dependent oxidoreductases, which catalyze the asymmetric reduction of an imine to the corresponding amine. In 2017, a subclass was identified, which is able to efficiently catalyze the direct reductive animation of a ketone to form the asymmetric amine, it was termed reductive aminases (RedAms) [32]. Chiral amines are of high interest for the pharmaceutical industry and thus, IREDs/RedAms were mentioned as an attractive emerging enzyme class in a review from the perspective of the pharmaceutical industry (GlaxoSmithKline-GSK) [33]. Shortly after that, GSK was the first company to enable a kilogram scale production of an API intermediate for the synthesis of the lysine-specific demethylase-1 inhibitor GSK-2879552 utilizing an engineered IRED (Section 2.3) [34]. Recently, the company Pfizer has described another process. In their work, they engineered a reductive amines for the synthesis of a key intermediate of abrocitinib (Figure 3), a Janus Kinase (JAK) inhibitor, which is a late-stage drug candidate for the treatment of atopic dermatitis [19]. After screening over 80 IREDs from their in-house enzyme library, they discovered _Sp_RedAm from _Streptomyces purpureus_ as being the most promising candidate but suffering from a low substrate tolerance.