7 research outputs found
The activation cycle of Rab GTPase Ypt32 reveals structural determinants of effector recruitment and GDI binding
Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/116366/1/feb2s0014579311007538-sup-m0005.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/116366/2/feb2s0014579311007538.pd
Mechanistic insights into the biosynthesis of polyketide antibiotics
Anthracyclines are a group of aromatic polyketide compounds with
significant medical importance due to their antineoplastic properties.
Doxorubicin and daunorubicin members of this family are among the two
most commonly used anticancer drugs. However, these compounds exhibit
severe side effects like cardiotoxicity and multi-drug resistance. A
promising approach towards the production of modified anthracyclines with
improved toxicity profiles appears to be combinatorial biosynthesis,
including the redesign of biosynthetic enzymes however, structural and
mechanistic information of the biosynthetic enzymes is necessary for the
redesigning approach.
The main focus of this thesis is the structural and functional studies of
several anthracycline biosynthetic enzymes. The crystal structures of two
polyketide cyclases, SnoaL and AknH were determined to 1.35Ă… and 1.9Ă…
respectively. These enzymes share very similar α+β structural folds and
catalyze a novel type of intramolecular aldol condensation reaction using
acid/base chemistry. Moreover, comparison of the cyclase structures,
followed by site-directed mutagenesis provided insights into the
structural basis of stereoselectivity of products in AknH.
Several enzymes from aromatic polyketide biosynthetic pathways, including
SnoaL/AknH provide illustrative examples of divergent evolution. The
related enzymes usually share the same fold but contain different
catalytic machineries to catalyze diverse reactions.
AknOx is a FAD-dependent oxidoreductase which is involved in the sugar
modification in aclacinomycins and catalyzes two consecutive oxidation
reactions. The structure of AknOx to 1.65Ă… resolution was obtained from a
pseudomerohedrally twinned crystal by MAD. AknOx crystals show an unusual
multi-domain twinning with six twin domains Investigation of the active
site and mutagenesis of the proposed residues revealed a unique feature
in AknOx compared to other flavoenzymes. The enzyme contains a dual
active site with two different sets of catalytic residues for catalyzing
two consecutive reactions