X-ray crystallographic studies on Particulate Methane Monooxygenase, Thioredoxin A and Arginine Decarboxylase
Author(s)
Andrell, Juni
Type
Thesis
Abstract
The work presented in this thesis describes the X-ray crystallographic studies of particulate
methane monoxygenase (pMMO) from Methylococcus capsulatus (Bath), thioredoxin A
(BsTrxA) from Bacillus subtilis and arginine decarboxylase (AdiA) from Escherichia coli.
1. pMMO is a respiratory enzyme that catalyses the first step in the metabolic pathway in
methanotrophic bacteria by converting methane to methanol. The crystal structure of this
integral membrane protein was determined by molecular replacement to 3.5 Å resolution.
The three metal sites in pMMO were confirmed to be a mononuclear copper site, a
dinuclear copper site and a mononuclear zinc site.
2. Thioredoxin is a ubiquitous protein present in nearly all known organisms. Its purpose
in the cell is to maintain cysteine-containing proteins in the reduced state by converting
intramolecular disulfide bonds to dithiols in a redox reaction. The crystal structure of an
active site mutant of BsTrxA was determined by molecular replacement to 1.5 Å
resolution. The structure shows a homodimer that resembles enzyme-substrate reaction
intermediates.
3. AdiA is a vitamin B6-dependent enzyme that catalyses the decarboxylation of arginine
into agmatine. It forms a part of an enzymatic system in E. coli that contribute to making
this organism acid resistant. The structure of arginine decarboxylase (AdiA) from E. coli
was determined by multiple isomorphous replacement and anomalous scattering (MIRAS)
methods to 2.4 Å resolution. The structure revealed a ~800 kDa decamer composed as a
pentamer of five homodimers. AdiA becomes active as the cellular environment becomes
more acidic. The structure of AdiA suggests how functional decamers associate with
decreasing pH or disassociates into inactive homodimers with increasing pH. The enzyme
mechanism and determinants for substrate specificity are discussed within the framework
of the structure and comparisons with related structures are made.
methane monoxygenase (pMMO) from Methylococcus capsulatus (Bath), thioredoxin A
(BsTrxA) from Bacillus subtilis and arginine decarboxylase (AdiA) from Escherichia coli.
1. pMMO is a respiratory enzyme that catalyses the first step in the metabolic pathway in
methanotrophic bacteria by converting methane to methanol. The crystal structure of this
integral membrane protein was determined by molecular replacement to 3.5 Å resolution.
The three metal sites in pMMO were confirmed to be a mononuclear copper site, a
dinuclear copper site and a mononuclear zinc site.
2. Thioredoxin is a ubiquitous protein present in nearly all known organisms. Its purpose
in the cell is to maintain cysteine-containing proteins in the reduced state by converting
intramolecular disulfide bonds to dithiols in a redox reaction. The crystal structure of an
active site mutant of BsTrxA was determined by molecular replacement to 1.5 Å
resolution. The structure shows a homodimer that resembles enzyme-substrate reaction
intermediates.
3. AdiA is a vitamin B6-dependent enzyme that catalyses the decarboxylation of arginine
into agmatine. It forms a part of an enzymatic system in E. coli that contribute to making
this organism acid resistant. The structure of arginine decarboxylase (AdiA) from E. coli
was determined by multiple isomorphous replacement and anomalous scattering (MIRAS)
methods to 2.4 Å resolution. The structure revealed a ~800 kDa decamer composed as a
pentamer of five homodimers. AdiA becomes active as the cellular environment becomes
more acidic. The structure of AdiA suggests how functional decamers associate with
decreasing pH or disassociates into inactive homodimers with increasing pH. The enzyme
mechanism and determinants for substrate specificity are discussed within the framework
of the structure and comparisons with related structures are made.
Date Issued
2008
Date Awarded
2008-12
Copyright Statement
Attribution NoDerivatives 4.0 International Licence (CC BY-ND)
Advisor
Carpenter, Liz
Iwata, So
Creator
Andrell, Juni
Publisher Department
Molecular Biosciences
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)