TY - JOUR
T1 - Structure and biocatalytic scope of thermophilic flavin-dependent halogenase and flavin reductase enzymes
AU - Menon, Binuraj R. K.
AU - Latham, Jonathan
AU - Dunstan, Mark S.
AU - Brandenburger, Eileen
AU - Klemstein, Ulrike
AU - Leys, David
AU - Karthikeyan, Chinnan
AU - Greaney, Michael F.
AU - Shepherd, Sarah A.
AU - Micklefield, Jason
N1 - Funding Information:
We acknowledge BBSRC (grant BB/K00199X/1), GlaxoSmithKline and CoEBio3 for financial support. We acknowledge Diamond Light Source for time on beamlines.
Publisher Copyright:
© The Royal Society of Chemistry 2016.
PY - 2016/10/21
Y1 - 2016/10/21
N2 - Flavin-dependent halogenase (Fl-Hal) enzymes have been shown to halogenate a range of synthetic as well as natural aromatic compounds. The exquisite regioselectively of Fl-Hal enzymes can provide halogenated building blocks which are inaccessible using standard halogenation chemistries. Consequently, Fl-Hal are potentially useful biocatalysts for the chemoenzymatic synthesis of pharmaceuticals and other valuable products, which are derived from haloaromatic precursors. However, the application of Fl-Hal enzymes, in vitro, has been hampered by their poor catalytic activity and lack of stability. To overcome these issues, we identified a thermophilic tryptophan halogenase (Th-Hal), which has significantly improved catalytic activity and stability, compared with other Fl-Hal characterised to date. When used in combination with a thermostable flavin reductase, Th-Hal can efficiently halogenate a number of aromatic substrates. X-ray crystal structures of Th-Hal, and the reductase partner (Th-Fre), provide insights into the factors that contribute to enzyme stability, which could guide the discovery and engineering of more robust and productive halogenase biocatalysts.
AB - Flavin-dependent halogenase (Fl-Hal) enzymes have been shown to halogenate a range of synthetic as well as natural aromatic compounds. The exquisite regioselectively of Fl-Hal enzymes can provide halogenated building blocks which are inaccessible using standard halogenation chemistries. Consequently, Fl-Hal are potentially useful biocatalysts for the chemoenzymatic synthesis of pharmaceuticals and other valuable products, which are derived from haloaromatic precursors. However, the application of Fl-Hal enzymes, in vitro, has been hampered by their poor catalytic activity and lack of stability. To overcome these issues, we identified a thermophilic tryptophan halogenase (Th-Hal), which has significantly improved catalytic activity and stability, compared with other Fl-Hal characterised to date. When used in combination with a thermostable flavin reductase, Th-Hal can efficiently halogenate a number of aromatic substrates. X-ray crystal structures of Th-Hal, and the reductase partner (Th-Fre), provide insights into the factors that contribute to enzyme stability, which could guide the discovery and engineering of more robust and productive halogenase biocatalysts.
KW - UKRI
KW - BBSRC
KW - BB/K00199X/1
UR - http://www.scopus.com/inward/record.url?scp=84990185741&partnerID=8YFLogxK
U2 - 10.1039/c6ob01861k
DO - 10.1039/c6ob01861k
M3 - Article
C2 - 27714222
AN - SCOPUS:84990185741
SN - 1477-0520
VL - 14
SP - 9354
EP - 9361
JO - Organic and Biomolecular Chemistry
JF - Organic and Biomolecular Chemistry
IS - 39
ER -