Research Opportunities 2010-11
Nelson, Berry and Warriner
Expanding the scope of biocatalysis through incorporation of unnatural
amino acids in an enzyme active site
Prof Adam Nelson, Prof Alan Berry and Dr Stuart Warriner
a.s.nelson@leeds.ac.uk , a.berry@leeds.ac.uk and S.L.Warriner@leeds.ac.uk
This proposal is representative of the projects currently on offer in our groups. For more details of active research
projects, please visit: www.asn.leeds.ac.uk , www.astbury.leeds.ac.uk/People/staffpage.php?StaffID=ABe and
http://www.chem.leeds.ac.uk/People/Warriner.html
The purpose of this project is to extend the scope of enzymatic catalysis through the incorporation of
unnatural amino acids into an enzyme active site. The position would suit a students with a strong
background in either chemistry or biochemistry, and a strong interest in chemical biology, biochemistry and
applications of enzymes in synthetic chemistry.
Enzymes are phenomenally powerful catalysts able to increase reaction rates by up to 1018-fold using the
diverse, but nonetheless limited, twenty natural amino acid side chains coupled with the ability to bind and
recruit metal ions or cofactors. We have previously used directed evolution to broaden the synthetic scope
of the Class I aldolase, sialic acid aldolase. For example, we discovered a variant enzyme (E192N) with
complementary substrate specificity,1 and we have created complementary enzymes for stereoselective
aldol reactions.2 Class I aldolases exploit enamine catalysis in which an catalytic lysine residue to form an
enamine intermediate, which then attacks the electrophile, the acceptor aldehyde (Panel A, Scheme). We
propose to extend the catalytic potential of sialic acid aldolase by using chemical methods to incorporate
unnatural residues in the active site. This approach could be used, for example, to allow iminium catalysis to
occur (Panel B, Scheme), an activation mode that is well known in organocatalysis but rare in enzyme
catalysis. Here, the modified residue would condense with an ,-unsaturated aldehyde to form an iminium
ion, and hence active the substrate towards nucleophilic attack. The overall approach could therefore allow
the catalytic activity of the aldolase enzyme to be dramatically expanded to allow, for example, the catalysis
of Michael or Diels-Alder reactions!
A. Enamine catalysis
R3
O
N
H
B. Iminium catalysis
Lysine in Class I aldolases
R4
R3
N
R2
R4
R2
reaction
with
R3
E
O
electrophile
R1
O
Incorporate unnatural
residue in place of lysine
R4
N
H
R3
N
R1
2
R
R2
R2
enamine
reaction 1
with R
 Michael reactions
HN
O
Pr 2N
O
OH
OH
CO2 H
R
2
iminium
 Aldol reaction catalysed by the E192N variant of sialic
acid aldolase
OH O
O
R4 nucleophile
Nu
OH
OH
Pr2N
O
indole
CO2 H
O
OH
OH
CO 2H
O
O
Pr 2N
O
Pr2 N
HO
CO2H
O
 Diels-Alder reaction
OH
OH
HO
HO
Pr2 N
O
H
O
CO 2H
This multidisciplinary project will provide opportunities for the student to receive training in synthetic
chemistry, protein chemistry and enzymology.
Please contact Professor Adam Nelson
(a.s.nelson@leeds.ac.uk) for further details about this opportunity.
References
1.
T. Woodhall, G. Williams, A. Berry and A. Nelson, “Creation of an aldolase for the parallel synthesis of sialic
acid mimetics”, Angew. Chem., Int. Ed., 2005, 44, 2109-2112.
2.
G. J. Williams, T. Woodhall, L. Farnsworth, A. Nelson and A. Berry, “Creation of a Pair of Stereochemically
Complementary Biocatalysts”, J. Am. Chem. Soc. 2006, 128, 16238-16247.
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