My laboratory uses approaches derived from both chemistry
and biology to investigate molecular mechanisms of cellular
biochemistry and physiology. Projects are organized around
the themes of purine/pyrimidine metabolism and protease
action and include problems in inhibitor design, protein
engineering and cellular metabolism. Most projects have
clinical relevance. Techniques vary with the particular
project, and can entail molecular biology, single molecule
experiments, organic synthesis, protein crystallography
and NMR spectroscopy as well as protein purification, enzyme
kinetics and mutagenesis. Some of our ongoing projects are
outlined below. For more information, please see the lab
web site.
Pathophysiological
mechanisms of retinal disease. Many inherited retinal
diseases are caused by mutations in proteins of the visual
cycle, which can easily explain why disease is photoreceptor-specific.
However, retinal disease can also result from mutations
in widely expressed proteins. The photoreceptor-specific
effects of these mutations are perplexing and pathophysiological
mechanisms are undefined. One such protein is inosine monophosphate
dehydrogenase type 1 (IMPDH1), which catalyzes a key step
in guanine nucleotide biosynthesis. The effects of the IMPDH1
mutations cannot be explained by the loss of enzyme activity.
We have recently discovered that IMPDH binds nucleic acids
and demonstrated that the disease-causing mutations perturb
nucleic acid binding. We are now investigating how this
defect causes the specific apoptosis of photoreceptor cells
with the aim of developing strategies for therapy.
Drug
Targets in Cryptosporidium parvum. This eukaryotic
parasite is an emerging opportunistic pathogen and potential
bio-warfare agent. The C. parvum oocyte is resistant
to the usual methods of water treatment, which has caused
spectacular outbreaks such as the infection of 40% of the
inhabitants of Milwaukee in 1993. C. parvum is resistant
to the usual antiparasitic drugs and currently used chemotherapy
is ineffective. We are mining the genome of C. parvum to reconstruct the parasite metabolic pathways and identify
new targets for drug development. Our biochemical characterization
of C. parvum IMP dehydrogenase suggests that this
enzyme is a promising target. We are currently developing
inhibitors specific for the parasite enzyme by screening
for novel lead compounds. We are also trying to determine
the structure of this enzyme with x-ray crystallography
that will enable the use of structure-based approaches for
drug design.
Dynamic structural determinants of drug selectivity. While
drug specificity is generally attributed simply to the topology
of the protein target, the contribution of dynamic processes
is becoming hard to ignore. We are investigating the dynamic
properties of inhibitor selectivity in two remarkable systems:
IMP dehydrogenase and serpins.
IMP dehydrogenase (IMPDH) is a target for immunosuppressive,
antiviral and anticancer chemotherapy. The enzyme undergoes
a large conformational change in mid-catalytic cycle that
is blocked by several drugs. Selectivity is determined by
how well the drug binds relative to the equilibrium of this
conformational change. This work involves detailed kinetic
characterization, structure determination and various biophysical
techniques to study protein dynamics.
Serpins
are ubiquitous inhibitors of serine proteases that regulate
blood coagulation, fibrinolysis and many other physiological
processes. Serpins react with their target protease to form
a stable adduct in a process that involves a 70 Å translocation
and structural rearrangements of both protease and inhibitor.
We are collaborating with Anne Gershenson to investigate
the mechanism of serpin inhibition with single molecule
experiments. This work builds on our classic experiments
delineating the structural determinants of specificity in
serine proteases.
Mechanistic enzymology. My laboratory has a long-standing
interest in the mechanisms of enzyme catalysis. The current
system of interest is the methionine salvage pathway, which
plays an important role in cell proliferation and involves
several challenging chemical transformations. The penultimate
step of this pathway is catalyzed by acireductone dioxygenase
(ARD). This remarkable protein catalyzes two different oxidative
cleavage reactions depending on the metal at the active
site. One of these reactions is part of the methionine salvage
pathway, and the other produces carbon monoxide, a gaseous
neurotransmitter. We are characterizing the human homolog
of ARD in collaboration with Tom Pochapsky.
Selected Publications:
IMP dehydrogenase: structure, mechanism, and inhibition. Hedstrom L. Chem Rev. 2009 Jul;109(7):2903-28.
Retinal isoforms of inosine 5'-monophosphate dehydrogenase type 1 are poor nucleic acid binding proteins. Xu D, Cobb G, Spellicy CJ, Bowne SJ, Daiger SP, Hedstrom L. Arch Biochem Biophys. 2008 Apr 15;472(2):100-4. [abstract]
An enzymatic atavist revealed in dual pathways for water activation. Min D, Josephine HR, Li H, Lakner C, MacPherson IS, Naylor GJ, et al. PLoS Biol. 2008;6(8):e206. [full text in PubMed Central] [abstract]
Targeting a prokaryotic protein in a eukaryotic pathogen: identification of lead compounds against cryptosporidiosis. Umejiego NN, Gollapalli D, Sharling L, Volftsun A, Lu J, Benjamin NN, et al. Chem Biol. 2008;15(1):70-7. [abstract]
A kinetic alignment of orthologous inosine-5'-monophosphate dehydrogenases. Riera TV, Wang W, Josephine HR, Hedstrom L. Biochemistry. 2008;47(33):8689-96. [abstract]
IMP dehydrogenase type 1 associates with polyribosomes translating rhodopsin mRNA. Mortimer SE, Xu D, McGrew D, Hamaguchi N, Lim HC, Bowne SJ, et al. J Biol Chem. 2008;283(52):36354-60. [full text in PubMed Central] [abstract]
IMP dehydrogenase-linked retinitis pigmentosa. Hedstrom L. Nucleosides Nucleotides Nucleic Acids. 2008;27(6):839-49. [abstract]
Short-lived protease serpin complexes: partial disruption of the rat trypsin active site. Liu L, Mushero N, Hedstrom L, Gershenson A. Protein Sci. 2007;16(11):2403-11. [abstract]
Regulation of nonproteolytic active site formation in plasminogen. Gladysheva IP, Sazonova IY, Houng A, Hedstrom L, Reed GL. Biochemistry. 2007;46(30):8879-87. [abstract]
Novel methylenephosphophosphonate analogues of mycophenolic adenine dinucleotide. Inhibition of inosine monophosphate dehydrogenase. Rejman D, Olesiak M, Chen L, Patterson SE, Wilson D, Jayaram HN, et al. J Med Chem. 2006;49(16):5018-22. [abstract]
Conformational distributions of protease-serpin complexes: a partially translocated complex. Liu L, Mushero N, Hedstrom L, Gershenson A. Biochemistry. 2006;45(36):10865-72. [abstract]
IMP dehydrogenase: structural schizophrenia and an unusual base. Hedstrom L, Gan L. Curr Opin Chem Biol. 2006;10(5):520-5. [abstract]
Spectrum and frequency of mutations in IMPDH1 associated with autosomal dominant retinitis pigmentosa and leber congenital amaurosis. Bowne SJ, Sullivan LS, Mortimer SE, Hedstrom L, Zhu J, Spellicy CJ, et al. Invest Ophthalmol Vis Sci. 2006;47(1):34-42. [full text in PubMed Central] [abstract]
View Complete Publication List on PubMed: Liz Hedstrom