Scientific Report 2008
Chemistry, Biology, and Inflammatory Disease
P. Wentworth, Jr., D. Angrish, J. Dambacher,
V. Dubrovskaya, R.K. Grover, J. Nieva, M. Puga, B.D. Song, M.M.R. Peram, J.K. Rogel,
S.R. Troseth, H. Wang, A.D. Wentworth
Our research is
interdisciplinary and involves aspects of bioorganic, biophysical, physical organic,
synthetic, and analytical chemistry coupled with biochemical techniques, cell-based
assays, and animal models. We are interested in uncovering new mechanisms of disease
in major conditions such as atherosclerosis, neurodegenerative diseases, ischemia-reperfusion
injury, macular degeneration, cancer, and infectious diseases.
Antibody-Catalyzed Water Oxidation Pathway
Our discovery that all antibody molecules
can catalyze the reaction between singlet oxygen and water to give hydrogen peroxide
is causing a revision of the idea that antibodies are only an adapter molecule within
the immune system, linking recognition and killing of foreign pathogens. We are
exploring both the chemical and biological aspects of this pathway, and new insights
into how the pathway plays a role in immune defense and inflammatory damage are
We are searching for the active site
for the antibody-catalyzed water oxidation pathway within the antibody structure.
We have cloned and expressed soluble individual domains (VHVL,
CH1CL, VH, VL, CH1, CL)
of the murine Fab 4C6. All of the domains can generate hydrogen peroxide when presented
with singlet dioxygen, suggesting that the driving force is related to the immunoglobulin
fold of the whole antibody.
INFLAMMATORY ALDEHDYES AND
We have shown that the inflammation-derived
cholesterol seco-sterols atheronal-A and atheronal-B trigger a deformation
in the secondary structure of the normally folded low-density lipoprotein apoB-100
into a proamyloidogenic form. In collaboration with J.W. Kelly and his group, Department
of Chemistry, we extended this model and showed that these cholesterol seco-sterols
also trigger the misfolding of amyloid β-peptide1-40,
leading to formation of fibrils similar to those observed in patients with Alzheimer's
disease. Using mutated synthetic sequences of amyloid β-peptide1-40,
we found that the accelerated aggregation of this protein only occurs when only
lysine 16, not lysine 28 or the N-terminal amino group of aspartic acid 1, of the
sequence is modified. More recently, in studies of inflammatory aldehyde—initiated
misfolding of antibody light chains (Bence-Jones proteins), we found that different
aldehydes can trigger different forms of aggregation in different proteins. Thus,
we have shown that the cholesterol seco-sterols atheronal-A and atheronal-B
accelerate an amorphous form of aggregation, whereas 4-hydroxynonenal induced an
amyloid form of aggregation of both λand κlight chains (Fig. 1).
Electron micrograph of fibrillar aggregation of antibody light chains induced by
cholesterol seco-sterol and 4-hydroxynonenal (shown in white).
Epidemiologic and clinical evidence point
to an increased risk of cancer when linked with chronic inflammation, in a process
thought to involve the establishment of a local inflammatory microenvironment conducive
to the development of neoplasia. However, because of the complex interrelationships
between the 2 conditions, the precise molecular and cellular mechanisms that underpin
this relationship remain largely unresolved.
found that the inflammation-derived cholesterol 5,6-seco-sterol aldehydes
atheronal-A and atheronal-B cause a loss of function of wild-type tumor suppressor
protein p53, the so-called guardian of the genome, in a process that involves p53
misfolding and amyloidogenesis.
Atheronal-A and atheronal-B, but not the aldehydes 4-hydroxynonenal and 4-hydroxyhexenal
derived from polyunsaturated fatty acids, induce misfolding of wild-type p53 into
an amyloidogenic form that binds thioflavin T and Congo red dye but cannot bind
to a consensus DNA sequence (Fig. 2). Treatment of lung carcinoma cells expressing
wild-type p53 with atheronal-A and atheronal-B leads to dysfunctional p53, as determined
by analysis of extracted nuclear protein and transcription activation of p21.
Optical microscopy images (100X) obtained with normal (upper) and cross-polarized
(lower) light of aggregates generated by incubation of hexahistidine-tagged native
p53 with atheronal-A and stained with Congo red.
results reveal a hitherto unknown chemical link between inflammation and cancer
and expand the already pivotal role of p53 dysfunction in the risk for cancer. The
increasing generality and specificity of aldehyde-initiated protein misfolding suggests
that inflammatory aldehydes and their posttranslational modification of amyloidogenic
peptides may be the chemical link between the known associations of inflammation,
oxidative damage, and various misfolding diseases.
INteraction Between Protozoan J-Binding Protein 1 And Glycosylated DNA
Current treatments of parasitic infections
such as leishmaniasis (cutaneous or visceral, Leishmania species), African
trypanosomiasis (sleeping sickness, Trypanosoma brucei), and American
trypanosomiasis (Chagas' disease, Trypanosoma cruzi) have limited
effectiveness, thereby increasing drug resistance and inherent toxic effects of
the drugs. Thus, an elucidation of new parasite-specific biological targets for
therapeutic agents is needed. In this regard, the discovery that DNA from members
of the order Kinetoplastida, but not other eukaryotes, contains an unusual modified
called base J, was a breakthrough. Extracts of several kinetoplastids contain a
J-binding protein (JBP) that specifically binds to J-containing duplex DNA. JBP-1
is essential in Leishmania.
As a drug target, JBP has merit. The
protein shares little homology with other proteins in the Protein Data Bank, and
it has a unique ligand, J-DNA containing telomeric stretches of double-stranded
DNA, that does not occur in other eukaryotes. However, a preliminary high-throughput
screen, focused on disrupting binding between JBP-1 and J-DNA, with a library of
compounds consisting of all the major drug pharmacophoric groups has revealed no
compounds of interest.
In parallel, we have studied the molecular
recognition that underlies JBP-1 recognition of glycosylated DNA. In collaboration
with D.P. Millar and D.A. Case, Department of Molecular Biology, we found that JBP-1
interacts with the J-containing DNA only when a critical conformation of the glucose
within the major groove is established. More recently, we discovered that low micromolar
concentrations of the DNA intercalators daunorubicin and mitoxantrone disrupt the
binding of JBP-1 with duplex DNA containing J-DNA. Modeling suggests that DNA binding
of the intercalators leads to distortion, which leads to disruption of the edge-on
conformation of the glucose within the major groove of the DNA.
Grover, R.K., Wentworth, P., Jr.
Emerging therapies for kinetoplastid diseases. Prog. Infect. Dis., in press.
Nieva, J., Shafton, A., Altobell,
L.J. III, Tripurenani, S., Rogel, J.K., Wentworth, A.D., Lerner, R.A., Wentworth,
P., Jr. Inflammatory aldehydes
accelerate antibody light chain amyloid and amorphous aggregation. Biochemistry
Scanlan, C.N., Ritchie, G.E., Baruah,
K., Crispin, M.D., Harvey, D.J., Singer, B.B., Lucka, L., Wormald, M.R., Wentworth,
P., Jr., Zitzmann, N., Rudd, P.M., Burton, D.R., Dwek, R.A. Inhibition
of mammalian glycan biosynthesis produces non-self antigens for a broadly neutralising,
HIV-1 specific antibody. J. Mol. Biol. 372:16, 2007.
Scheinost, J.C., Boldt, G.E., Wentworth,
P., Jr. Protein misfolding
diseases. In: Encyclopedia of Chemical Biology, Wiley Blackwell, New York,
Wang, H., Boldt, G.E., Offer, J., Wentworth, P., Jr.
Cholesterol seco-sterol-induced aggregation of methylated amyloid-β
peptides, insights into aldehyde-initiated fibrillization of amyloid-β .
Angew. Chem. Int. Ed. 47:3919, 2008.
Temperini, C., Cecchi, A., Boyle,
N.A., Scozzafava, A., Cabeza, J.E., Wentworth, P., Jr., Blackburn, G.M., Supuran,
C.T . Carbonic anhydrase inhibitors.
Interaction of 2-N,N-dimethylamino-1,3,4-thiadiazole-5-methylsulfonamide
with 12 mammalian isoforms: kinetic and x-ray crystallographic studies. Bioorg.
Med. Chem. Lett. 18:999, 2008.
Wentworth, P., Jr., Witter, D.
Antibody-catalyzed water-oxidation pathway. Pure Appl. Chem. 80:1849, 2008.