Angewandte
Chemie
DOI: 10.1002/anie.201107695
Enzyme Catalysis
Tetrahydroxynaphthalene Reductase: Catalytic Properties of an
Enzyme Involved in Reductive Asymmetric Naphthol
Dearomatization**
Michael A. Schꢀtzle, Stephan Flemming, Syed Masood Husain, Michael Richter, Stefan Gꢁnther,
and Michael Mꢁller*
Alcohol dehydrogenases are mainly applied for the reduction
of aliphatic ketones. By contrast, a biosynthetic point of view
hints at broader catalytic activities. For example, the metab-
olism of aromatic compounds in fungi and bacteria proceeds
through an aerobic or anaerobic route. While mono- or
dioxygenases are involved in the aerobic pathways, oxidor-
eductases are required for the anaerobic metabolism, thus
leading to the formation of acetyl coenzyme A. Additionally,
polyphenolic polyketide synthase (PKS) products degrade by
a reduction–dehydration sequence during secondary metab-
olite synthesis.[1] This deoxygenation strategy was found as
Scheme 1. Fungal DHN-melanin biosynthesis; dashed box: prevalent
keto–enol tautomerism of 2 in solution.[2–6,15–21]
a key step in 1,8-dihydroxynaphthalene (DHN, 1)-melanin
biosynthesis of various fungi[2–6] and is also proposed for the
formation of aflatoxin,[7,8] actinorhodin,[9] and chrysopha-
nol.[10,11] Furthermore, polyhydroxynaphthalenes represent
branching points in several secondary metabolite synthe-
ses.[12–14] A high degree of metabolic diversity can be found,
especially in the group of spirodioxynaphthalenes.[12]
steps are believed to take place via the 3-keto tautomers of 2
and 4.[20,21] T4- and T3HNR of Magnaporthe grisea involved in
this route show 46% sequence identity and exhibit a prefer-
ence for T4HN (2) and T3HN (4), respectively.[15] The two
reductases are members of the short-chain dehydrogenase/
reductase (SDR) family. This family is known for sharing the
same structural motif, though the members exhibit few
sequence similarities.[15,19,22–25] The common conserved motif
is the core structure for divergent reductases, but also
represents the backbone for a catalytic-promiscuous family.[26]
Dearomatization strategies represent a powerful and
direct approach to cyclic building blocks in natural product
synthesis.[27,28] However, dearomatization reactions concur-
rent with asymmetric catalysis in one step are very challeng-
ing.[27] Thus, T4HNR and T3HNR may represent valuable
tools for catalytic, asymmetric dearomatization.
Herein, we focus on the structure–activity relationship of
T4HNR to gain insights into the dynamics of its catalytic cycle.
A broad substrate range allowed for the identification of an
essential structural motif of naphtholic substrates and of
major active-site interactions. The mutational effect of C-
terminal truncation agrees with the concept of stabilization by
the C-terminal carboxylate as an explanation for the substrate
preference of T4HNR. This structural feature may help to find
further naphthol reductases.
DHN (1), as the monomeric unit of DHN-melanin, is
produced by means of
a
double deoxygenation
(Scheme 1).[2–6] The PKS product 1,3,6,8-tetrahydroxynaph-
thalene (T4HN, 2) is reduced by tetrahydroxynaphthalene
reductase (T4HNR) to scytalone (3),[15] which is readily
dehydrated by scytalone dehydratase (SD) to 1,3,8-trihydr-
oxynaphthalene (T3HN, 4).[16–18] In the same manner, 4 is
reduced by trihydroxynaphthalene reductase (T3HNR) to
vermelone (5) and aromatized by SD to 1.[19] The reduction
[*] M. A. Schꢀtzle, S. Flemming, Dr. S. M. Husain, Dr. M. Richter,
Dr. S. Gꢁnther, Prof. Dr. M. Mꢁller
Institut fꢁr Pharmazeutische Wissenschaften
Albert-Ludwigs-Universitꢀt Freiburg
Albertstrasse 25, 79104 Freiburg (Germany)
E-mail: michael.mueller@pharmazie.uni-freiburg.de
Dr. M. Richter
Laboratory for Biomaterials, Empa–Swiss Federal Laboratories for
Materials Science and Technology
Lerchenfeldstrasse 5, 9014 St. Gallen (Switzerland)
[**] Financial support of this work by the Deutsche Forschungsge-
meinschaft (IRTG 1038) is gratefully acknowledged. We are grateful
to E. Breitling for skillful technical support, to V. Brecht for
T4HNR was cloned and expressed using a slightly modi-
fied method of Thompson et al.[15] and the cell extract was
used without further purification. Despite its susceptibility to
oxidation by air, the physiological substrate 2 gave a con-
version of 47% with 33% yield of scytalone (3) being
obtained. The absolute configuration was determined by CD
spectroscopy to be R, and the enantiomeric excess (ee) was
found to be > 95% (Table 1 and the Supporting Information).
measurement of NMR spectra, to Dr. S. Lꢁdeke for VCD measure-
ment, and to Prof. Dr. U. Kꢁck and Prof. Dr. F. J. Leeper for helpful
discussions. We acknowledge the use of the computing resources
ˇ
provided by the Black Forest Grid initiative and T. L. Rizner for
providing 17b-HSDcl.
Supporting information for this article is available on the WWW
Angew. Chem. Int. Ed. 2012, 51, 2643 –2646
ꢀ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
2643