Asymmetric synthesis of primary amines catalyzed by thermotolerant fungal reductive aminases

Article abstract of DOI:10.1039/d0sc02253e

Chiral primary amines are important intermediates in the synthesis of pharmaceutical compounds. Fungal reductive aminases (RedAms) are NADPH-dependent dehydrogenases that catalyse reductive amination of a range of ketones with short-chain primary amines supplied in an equimolar ratio to give corresponding secondary amines. Herein we describe structural and biochemical characterisation as well as synthetic applications of two RedAms fromNeosartoryaspp. (NfRedAm andNfisRedAm) that display a distinctive activity amongst fungal RedAms, namely a superior ability to use ammonia as the amine partner. Using these enzymes, we demonstrate the synthesis of a broad range of primary amines, with conversions up to >97% and excellent enantiomeric excess. Temperature dependent studies showed that these homologues also possess greater thermal stability compared to other enzymes within this family. Their synthetic applicability is further demonstrated by the production of several primary and secondary amines with turnover numbers (TN) up to 14 000 as well as continous flow reactions, obtaining chiral amines such as (R)-2-aminohexane in space time yields up to 8.1 g L^?1h^?1. The remarkable features ofNfRedAmand NfisRedAm highlight their potential for wider synthetic application as well as expanding the biocatalytic toolbox available for chiral amine synthesis.

Full text of DOI:10.1039/d0sc02253e

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Asymmetric synthesis of primary amines catalyzed
by thermotolerant fungal reductive aminases†
Cite this: DOI: 10.1039/d0sc02253e
a
b
a
All publication charges for this article Juan Mangas-Sanchez,
Mahima Sharma,
Sebastian C. Cosgrove,
have been paid for by the Royal Society
of Chemistry
a
a
a
a
Jeremy I. Ramsden, James R. Marshall, Thomas W. Thorpe, Ryan B. Palmer,
Gideon Grogan* and Nicholas J. Turner
b
a
*
Chiral primary amines are important intermediates in the synthesis of pharmaceutical compounds. Fungal
reductive aminases (RedAms) are NADPH-dependent dehydrogenases that catalyse reductive amination of
a range of ketones with short-chain primary amines supplied in an equimolar ratio to give corresponding
secondary amines. Herein we describe structural and biochemical characterisation as well as synthetic
applications of two RedAms from Neosartorya spp. (NfRedAm and NfisRedAm) that display a distinctive
activity amongst fungal RedAms, namely a superior ability to use ammonia as the amine partner. Using
these enzymes, we demonstrate the synthesis of a broad range of primary amines, with conversions up
to >97% and excellent enantiomeric excess. Temperature dependent studies showed that these
homologues also possess greater thermal stability compared to other enzymes within this family. Their
synthetic applicability is further demonstrated by the production of several primary and secondary
Received 21st April 2020
Accepted 30th April 2020
amines with turnover numbers (TN) up to 14 000 as well as continous flow reactions, obtaining chiral
À1 À1
amines such as (R)-2-aminohexane in space time yields up to 8.1 g L
h . The remarkable features of
DOI: 10.1039/d0sc02253e
NfRedAm and NfisRedAm highlight their potential for wider synthetic application as well as expanding
rsc.li/chemical-science
the biocatalytic toolbox available for chiral amine synthesis.
10
compounds and small amine donors in equimolar quantities.
We have recently demonstrated that these enzymes catalyze the
Introduction
Chiral amines are a privileged functional group, ubiquitous in condensation of cyclic and aliphatic ketones with short-chain
natural products, pharmaceuticals, agrochemicals and ne amines as well as the subsequent reduction of the imine inter-
1
11
chemicals. Consequently, the discovery and development of mediate, although the substrate scope has until now been
more efficient ways to synthesise chiral amines continues to be mainly limited to the preparation of secondary and tertiary
a major research topic in synthetic chemistry. In this context, amines. The asymmetric synthesis of chiral primary amines,
biocatalysis offers an alternative to conventional chemical strat- synthesized via reductive amination of ketones using inexpensive
12
egies that oen rely on the use of hazardous reagents and reac- ammonia, continues to be a challenge in synthetic chemistry.
tion conditions. Over the last 20 years, a substantial effort has Biocatalytic strategies for the asymmetric synthesis of chiral
been made in the eld and a signicant number of biocatalysts primary amines include the application of transaminases (TAs),
2–6
are now available for this task. Among these enzymes, imine which catalyse the pyridoxal phosphate-dependent amination of
3
,13
reductases (IREDs), which catalyze the NAD(P)H-dependent ketone at the expense of an ammonia donor,
reduction of prochiral imines to chiral amines, have assumed dehydrogenases (AmDHs),
and amine
which catalyse the NAD(P)H-
an important role within the biocatalytic toolbox for chiral amine dependent reductive amination of ketones to primary amines
5,14–16
7–9
synthesis, following their discovery by Mitsukura et al. A sub- (Fig. 1). Nevertheless, these two families of enzymes present some
group of IREDs, termed reductive aminases (RedAms), difficulties with respect to their application. In TA-catalysed
possesses additionally the distinctive ability to carry out reductive processes, in addition to the need of an amine donor in high
aminations in water, when presented with a range of carbonyl concentration, such as alanine or isopropylamine, equilibrium
issues make the use of enzymatic cascades or product removal
1
7–21
strategies mandatory to achieve high yields.
AmDHs are
a
School of Chemistry, University of Manchester, Manchester Institute of Biotechnology,
31 Princess Street, Manchester, M1 7DN, UK. E-mail: nicholas.turner@manchester.
engineered amino acid dehydrogenases (AADHs), which use
ammonium salts as amine partners and constitute an emerging
1
ac.uk
b
York Structural Biology Laboratory, Department of Chemistry, University of York, source of biocatalysts, however their substrate scope is still
14,15,22–25
YO10 5DD York, UK
limited.
A new family of natural AmDHs (nat-AmDHs) has
†
Electronic supplementary information (ESI) available. See DOI:
0.1039/d0sc02253e
been recently described that will bring diversity and therefore
1
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Reductive amination with primary amines
The substrate scope of NfRedAm and NsRedAm was initially
assessed by combining a series of carbonyl compounds (1–7)
with 1–5 equivalents of different primary amines (Fig. S4 in the
ESI†). As previously observed with other fungal RedAms, the
best results were observed aer combining ketoesters, cyclic
and aliphatic ketones such as ethyl levulinate (1), cyclohexa-
none (3) or 2-hexanone (4), with short-chain amines with certain
p-character such as propargylamine (a), allylamine (b) and
cyclopropylamine (c). This permits access to a wide range of
chiral secondary amines, lactams and amino esters in good to
excellent conversions (70% to >97%) and moderate to good
enantiomeric excess and at low equivalents of the amine donor.
Secondary amines derived from 2-octanone (6), 2-decanone (7)
and 4-phenyl-2-butanone (5) can also be synthesised in this
manner as well as d-amino esters (2a,b) in moderate to good
conversions (45–70%). The acceptance of benzylamine, which
has previously been reported for an imine reductase homologue
Fig. 1 Biocatalytic reductive amination of ketones. (a) Reductive
aminations mediated by amine dehydrogenases (AmDHs) using
ammonia. (b) Formal reductive amination catalysed by transaminases
(
(
TAs) using isopropylamine as amine donor. (c) Reductive aminase
RedAm) catalysed reductive aminations with primary amines. (d)
30
RedAm catalysed reductive aminations with ammonia.
from Myxococcus stipitatus, was also explored. Remarkably,
one equivalent of benzylamine (d) with 3 yielded 85% of the
corresponding secondary amine 3d in 85% conversion for
NfRedAm, whereas only traces of the product were observed for
NsRedAm. NfRedAm also outperforms other RedAms such as
enable the expansion of the substrate scope although to-date it
2
6,27
still remains rather limited.
IREDs have also been used to
28
access primary amines, although poor conversions were ob-
tained for the amination of prochiral ketones. In this context, and
given that we have previously observed that, by forcing reaction
conditions, RedAms can also perform the reductive amination of
11
AdRedAm, where only 26% conversion was obtained. In terms
of stereoselectivity, enantiomeric excesses ranged from poor to
excellent for both RedAms and are notably different or even
enantio-complementary in some instances (as seen in the case
of ketoester 1–2, Fig. S3 in the ESI†). Remarkably, both the
enantiopreference and extent of selectivity appeared to be both
ketone- and amine-dependent, an observation which, given the
sequence similarity between both enzymes (94% sequence ID),
stresses the complexity of the mechanisms that govern the
stereoselectivity of these reactions.
10
a wider range of ketones and amines, we envisaged that
homologues within this family could also accept a broader range
of carbonyl compounds as well as ammonia to access primary
amines. In a recent report, one of our groups described the ability
of two of these homologues – NfRedAm from Neosartorya fumigata
and NsRedAm from Neosartorya scheri – for the reduction of the
imine formed between acetophenone and ammonia, albeit at
concentrations of 5 mM and 250 mM respectively, to give
conversions to the primary amine product which, although
Amination using ammonia
11
modest, were superior to previously described RedAms such as Following our preliminary observation, the initial assessment
29
AdRedAm from Ajellomyces dermatitidis. In this study we exploit of the activity of NfRedAm and NsRedAm with a range of
this initial observation in the characterization of the activity of substrates conrmed that these enzymes appeared to display
NfRedAm and NsRedAm and their application to the scalable superior activity when using ammonia (f) as a donor compared
synthesis of primary and secondary amines using ammonia.
to previously studied RedAms such as AtRedAm and AdRedAm.
For instance, with cyclohexanone (3), when using 5 equivalents
of ammonia (f), 47% conversion to cyclohexylamine (4f) was
observed in the AdRedAm-catalysed process, whereas 90% and
81% conversions were obtained for NfRedAm and NsRedAm,
respectively. Similarly, 14% conversion was obtained for AdRe-
dAm in the reductive amination of 2-hexanone (4) and
ammonia (f), whilst a 72% conversion was observed for NfRe-
Results and discussion
The two target enzymes in this study, NfRedAm from Neo- dAm under the same reaction conditions. This distinctive
sartorya fumigatus and NsRedAm from Neosartorya scheri, activity was further explored by screening these RedAms across
each displayed excellent levels of expression in E. coli a panel of structurally diverse ketones 3–20 (Fig. 2) using 1 M
BL21(DE3). The initial report, suggesting that these enzymes ammonia (100 equivalents) to mimic the optimal conditions in
15
may display some activity for the reduction of the imine the AmDH-catalysed processes. Considering the possibility
resulting from the ambient reaction of acetophenone and that the cofactor regeneration system could also be affected
ammonia, prompted an in-depth investigation of the amination under such challenging reaction conditions, different NADPH
of ketones using NfRedAm and NsRedAm for the synthesis of regeneration strategies (glucose dehydrogenase from Bacillus
primary amines.
subtilis, CDX-901 glucose dehydrogenase from Codexis and
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12f with 96% ee, 4-nitroacetophenone (9), 4-uoro-
acetophenone (11) or 4-phenyl-2-butanone (5) gave moderate
to good conversions (44–76%) yielding the corresponding (R)-
amine products 5f, 8f, 9f and 11f in moderate to excellent
enantiomeric excess (40–96%). Other aromatic substrates such
as 1-tetralone (10), 3-acetyl pyridine (12), 2-acetyl pyridine (13),
1-indanone (14) and propiophenone (16), low to poor conver-
sions were obtained (up to 16%) although in high enantiomeric
excess (>93%). In no cases were the alcohol products observed.
Structure of NfRedAm
The structure of NfRedAm was determined in two forms: an apo-
˚
form, at a resolution of 2.25 A, in space group P3 21 with one
2
molecule in the asymmetric unit (asu), and a complex with the
+
NADP cofactor, in space group P1, at a lower resolution of 2.77
A˚ , with eight molecules in the asu forming four dimers.‡
However, size-exclusion chromatography-multiangle light scat-
tering (SEC-MALS) analysis conrmed that NfRedAm exists as
a dimer in solution (Fig. S3 in the ESI†). The structure was
solved by molecular replacement using the structure of AtRe-
Fig.
access chiral primary amines. Conditions: 10 mM ketone in 100 mM pH
Tris–HCl buffer containing 10 mM (green) or 1 M (black) NH Cl, 1 mM
2
RedAm-catalysed reductive amination using ammonia to
11
dAm (PDB 6EOD, 55% sequence identity) as a model. Data
collection and renement statistics can be found in Table S2 in
9
4
+
À1
NADP , 40 mM glucose and 0.5 mg mL CDX-GDH. Conversions the ESI.† The structure of the dimeric form of the enzyme is
determined after 24 h based on the areas of substrate and product by shown in Fig. 3A. The structure conforms to the canonical
GC-FID.
model determined for IRED-family enzymes, featuring two
monomers, each composed of an N-terminal Rossmann
domain, connected to a C-terminal helical bundle by a long
a variant of the phosphite dehydrogenase from Pseudomonas
alpha helix. The dimer is formed through reciprocal domain
sharing, with the active sites formed at the interface between
the N-terminal domain of one subunit, and the C-terminal
28
stutzeri (PtDH) ) and different ammonium sources [NH
NH SO and NH OH] were evaluated (Fig. S5–S7 in the ESI†).
For our reactions, GDH (CDX-901)/Glucose as NADPH regener-
4
Cl,
(
4
)
2
4
4
31
domain of the other. Analysis using the DALI server shows
that the monomer superimposed with its closest homologues –
the RedAms from Aspergillus oryzae (PDB 5G6R; 58% sequence
identity) and Aspergillus terreus (5OJL; 57%) with rmsd values of
ation system and 1 M NH Cl in 100 mM Tris–HCl pH 9 were
4
found to be the best conditions.
As previously observed for the reductive amination with
1
0,11
primary amines,
the best results were obtained for cyclic and
Cl, full conversions were found for
˚
1
.3 and 1.8 A over 289 and 284 C-alpha atoms respectively.
aliphatic ketones. At 1 M NH
4
The structure of the active site is shown in Fig. 3B. The
cyclic ketones such as 3, 15 and 18. Cyclohexen-2-one (19) was
also accepted affording cyclohex-2-en-1-amine (19f) as a sole
product in up to 87% conversions. The corresponding saturated
product cyclohexylamine (3f) was not detected in any case. For
aliphatic substrates, moderate to excellent conversions were
afforded although a drop in activity was observed with longer
substrates. Remarkably, a switch in stereoselectivity was found
in this series as well. (R)-2-Aminohexane (4f) can be obtained in
full conversion and >97% ee. A 75% conversion was observed in
the reductive amination of 6 to afford (R)-2-aminooctane (6f) in
majority of active site residues are conserved between NfRedAm
and AspRedAm, including D167 (NfRedAm numbering) and
Y175, thought to be involved in the activation of small amine
nucleophiles and in securing the carbonyl group of the ketone
substrate respectively. T120 in NfRedAm replaces V121 in
+
AspRedAm behind the NADP cofactor as viewed in Fig. 3B.
With many active site residues conserved, the reasons for the
improved reduction of primary imines was not clear. In order to
further investigate the comparative biophysical characteristics
of the RedAm enzymes we determined the melting temperature
of AspRedAm, NfRedAm, NsRedAm using circular dichroism
34% ee and a switch to the S-product was observed for 2-dec-
anone (7), obtaining (S)-2-aminodecane (7f) in 41% conversion
and 84% ee. Other aliphatic substrates such as cyclohexyl
methyl ketone (17) and 3-octanone (20) can also be aminated in
this manner in moderate to good conversions (49–92%). For 17,
a switch in the general stereoselectivity was again observed,
obtaining (S)-17f in 14% and 72% ee for NfRedAm and NsRe-
dAm respectively. Aromatic ketones could also be aminated
(
Fig. 4).
The unfolding of the enzymes was studied by recording
ꢀ
CD222 at different temperatures (20–85 C) as a measure of
thermodynamic stability (Fig. 4). Importantly, the melting
temperatures (T
higher than AspRedAm, which demonstrated the lowest Tm
value amongst these fungal RedAms.
ꢀ
m
) of the homologues were found to be 6–10 C
29
under these conditions. As previously described, NfRedAm
catalyzed the reductive amination of acetophenone (8) to amine
We also evaluated the thermostability of NfRedAm compared
to AspRedAm by measuring the residual activity of both
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+
Fig. 3 (A) Structure of dimer of NfRedAm in ribbon format with subunits shown in blue and pink. (B) Detail of active site of NfRedAm with NADP
shown in cylinder format with carbon atoms in grey.
selected as a control reaction as both enzymes display high
activity for this transformation. Aer an incubation time of only
20 min, a 50% decrease in activity was observed for AspRedAm
whilst a slight drop was found for NfRedAm. Remarkably,
NfRedAm does not seem to be signicantly affected by the
thermal treatment and retains most of its reductive amination
ꢀ
activity aer 1 h at 50 C.
Synthetic potential for large-scale processes
32
Given the potential of RedAms for large-scale processes, we
used 4-phenyl cyclohexanone (15) and ammonia (f) as model
reaction components to test higher substrate loadings (Fig. S8
and S9 in the ESI†). In this case, turnover numbers (TN) up to
À1
1
400 were obtained at 40 mM concentration (7 g L ) although
Fig. 4 Thermal denaturation studies as a measure of thermodynamic
stability. (a) CD spectra of NfRedAm as the temperature is increased this system is restricted by the limited solubility of the starting
ꢀ
from 20 to 85 C. (b) Melting temperatures of AspRedAm, NfRedAm
material and further reaction engineering would be necessary to
reach higher productivities. With conversion at 97%, a space
and NfisRedAm. CD220 plotted against temperature to calculate the
melting temperatures of (c) AspRedAm and (d) NfRedAm and
À1 À1
time yield (STY) of 0.28 g L
catalyst productivity of 6.8 gprod
h
g
was obtained, and a bio-
.
NfisRedAm.
enzÀ1
In the reductive amination of 2-hexanone (4), TN up to 3250
ꢀ
enzymes aer incubation at 50 C at different times (Fig. 5). In
this case, the reductive amination of cyclohexanone (3) and
À1
were obtained at 80 mM substrate concentration (8 g L ). This
À1 À1
represented a low STY of only 0.34 g L
h , but with biocatalyst
ꢀ
allylamine (b) at equimolar concentrations and 30 C was
À1
loading at only 0.5 mg mL it delivered a reasonable produc-
À1
tivity of 16 gprod
g
enz . However, TN decreased at higher
concentrations and additional DMSO loadings to increase the
solubility of the starting material led to lower productivities. The
use of continuous ow was investigated to try to improve the
33
process conditions for these enzymes. More specically,
immobilisation of NfRedAm was investigated as the immobilised
preparations are oen stabilised by the solid supports, and can
34
be applied in continuous packed-bed reactors. We applied EziG,
a porous silica-based affinity resin, as we had demonstrated
35–37
previously that it could stabilise AspRedAm.
NfRedAm was
immobilised and tested under several conditions with 2-hex-
anone (4) (Section S6 in the ESI†). The optimal conditions
permitted the continuous production of (R)-2-aminohexane ((R)-
Fig. 5 Relative activity of AspRedAm (black) and NfRedAm (green) in
ꢀ
the equimolar bio-reductive amination of 3 and b at 30 C after
ꢀ
4f) for up to 12 h. The optimised conditions (see ESI Fig. S12,
incubation at 50 C at different times. 100% activity corresponds to
1
NfRedAm.
À1 À1
.03 Æ 0.03 U mgÀ1 for AspRedAm and 0.77 Æ 0.11 U mg for
À1
entry 6†) delivered a STY of 8.1 g L
h , which is much higher
than the equivalent batch reaction. The biocatalyst productivity
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enzÀ1
, with TN up to 1400. Isolation issues must be
was 4.86 gprod
g
Experimental
Gene expression and protein purication
overcome to allow this to be applied on a larger scale, however
this initial proof-of-concept study demonstrates the versatility of
these biocatalysts for process intensication (see ESI† for full The plasmids containing the genes for target enzymes were
details of ow experiments). We also tested these enzymes for used to transform E. coli BL21(DE3) competent cells for gene
reductive amination with primary amines. At 100 mM 3 and 2 expression. Pre-cultures were grown in LB-medium (5 mL)
À1
ꢀ
equivalents of methylamine, TN of 2900 and 3400 were obtained containing 30 mg mL
kanamycin for 18 h at 37 C with
for NfRedAm and NsRedAm, respectively. At higher substrate shaking at 180 rpm. 0.6 L volume cultures were inoculated with
ꢀ
concentrations, NfRedAm displayed up to 14 000 TN in the the pre-culture (6 mL) and incubated at 37 C, with shaking at
equimolar reductive amination of 3 and b. In the reaction of 5 180 rpm until an OD600 of 0.6–0.8 was reached. Gene expression
(
100 mM) and cyclopropylamine (400 mM), TN around 2000 were was induced by addition of IPTG (1 mM) and shaking was
ꢀ
obtained whilst remarkably, for 4, full conversions were continued overnight at 20 C with shaking at 180 rpm. The cells
observed. We reduced the amine concentration to 200 mM (2 eq.) were then harvested by centrifugation at 5000g for 20 min and
observing no decrease in conversion. 4 was also tested at 200 mM resuspended in 50 mM Tris–HCl buffer pH 7.5, containing
(
400 mM c) observing TN up to 6800 with NfRedAm. Encouraged 300 mM NaCl. Cells were disrupted by ultrasonication for 3 Â
by these results, we also looked at other combinations beyond 5 min, with 30 s on and 30 s off cycles, and the suspension was
equimolar ratios of both partners (Fig. S10 in the ESI†). In centrifuged at 50 000g for 30 min to yield a clear lysate. The N-
particular, we focused on acetophenone (8) and 1-(4- terminal His6-tagged proteins were puried using immobilised-
uorophenyl)propan-2-one (11), given that secondary amines metal affinity chromatography (IMAC) using Ni-NTA column.
containing these scaffolds are valuable intermediates in different
industries, and a–c as amine partners. For 8, cyclopropylamine RedAm-catalysed bio-reductive amination
(
5
c) was found to be the best amine partner with conversions up to
6% with NsRedAm. For a and b as amine donors, NfRedAm
performed better although moderate conversions were achieved
21 and 25%, respectively). For 11, b was found to be the best
A 500 mL reaction mixture contained 20 mM D-glucose, 0.5 mg
mL GDH (Codexis, CDX-901), 0.5 mM NADP , 1 mg mL
À1
+
À1
puried RedAm, 10 mM ketone, in 1 M NH
4
Cl pH 9 NaPi buffer
(
ꢀ
adjusted and 2% v/v DMSO. Reactions were incubated at 30 C
with 250 rpm shaking for 24 h, aer which they were quenched
by the addition of 30 mL of 10 M NaOH and extracted twice with
nucleophilic partner, observing full conversion to the corre-
sponding chiral secondary amine 11b for both NfRedAm and
NsRedAm. In all cases, excellent enantiomeric ratios were
detected (from 91 to >97%). These results constitute a promising
starting point and show the synthetic versatility and potential of
these enzymes for industrial biocatalysis.
500 mL tert-butyl methyl ether. The organic fractions were
combined and dried over anhydrous MgSO and analysed by
4
HPLC or GC-FID on a chiral stationary phase.
Conflicts of interest
The authors declare no conicts of interest.
Conclusions
Acknowledgements
The activity of two enzymes from the fungal reductive aminase
J. M.-S. and M. S. were funded by grant BB/M006832/1 from the
UK Biotechnology and Biological Sciences Research Council. N.
J. T. is grateful to the ERC for the award of an Advanced Grant
(RedAm) clade, NfRedAm and NsRedAm, has been charac-
terised. While they possess a similar substrate scope to previously
described RedAms in the reductive amination with primary
amines, their ability to enable the reductive amination of ketones
using ammonia was found to be much higher. This distinctive
activity was further explored showing a broad substrate scope,
which highlights the versatility of these enzymes to serve as
scaffolds for evolution towards t-to-purpose biocatalysts in
industrial processes. These RedAms also display a greater ther-
mostability compared to other enzymes within the same family.
Finally, an initial assessment of the industrial potential of
(Grant number 742987). S. C. C. would like to acknowledge the
EPSRC UK Catalysis Hub for funding. We also thank Dr James
Galman from the Manchester Institute of Biotechnology for the
plasmid containing the phosphite dehydrogenase variant from
Pseudomonas stutzeri (PtDH).
Notes and references
+
‡
The coordinates for NfRedAm-apo and NfRedAm-NADP complex were depos-
NfRedAm and NsRedAm was carried out by performing reac- ited in the PDB with accession codes 6SKX and 6SLE respectively.
tions at high substrate loadings of selected substrates obtaining
TN up to 14 000, as well as reactions at higher amine-to-ketone
ratios to expand their synthetic operational landscape. Reac-
tions in continuous ow were also performed, optimising the
1 C. Wang and J. Xiao, in Stereoselective Formation of Amines,
ed. C. Wang and J. Xiao, Springer-Verlag Berlin Heidelberg,
2014, pp. 261–282.
system for the production of (R)-2-aminohexane with a STY of
2 V. F. Batista, J. Galman, D. C. Gouveia Alves Pinto,
A. M. S. Silva and N. J. Turner, ACS Catal., 2018, 8, 11889–
11907.
À1 À1
8.1 g L
h , which show the extraordinary capabilities and
synthetic potential of these biocatalysts for large-scale use.
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