Senior Projects 2010 | St. Lawrence University Chemistry

Senior Projects 2010

Chemistry majors graduating in 2010 include Kristin Berretta, Dan DePuccio, Matthew DePuccio, Michelle Dumas, Josh Johnson, Connor Kellogg, Ryan Littlefield, Chris Lundberg, Geoff Millard, Brian Palmer, Alyssa Pirinelli (at right), Hillary St. John, Tim Senter, Marcus Tuttle and Cathy Yancey.

Biochemistry majors graduating in 2010 include Samantha Berghorn, Brian Brownell Jr., Matt DePuccio, Michelle Dumas, Luke Grover, Peterson Maina and Brian Palmer.  In addition, Reed Carr is a senior Environmental Studies-Chemistry major. 

What are our seniors researching?

Kristin
Berretta
: Chemical Models of Anti-Cancer Pharmaceuticals Based on
Ruthenium Compounds with Organic Side Chains (with Dr. Samantha
Glazier)

Samantha
Berghorn
: BMH1 and BMH2 (with Dr. Emily Dixon)

Brian Brownell: 14-3-3 Proteins:
BMH1 and BMH2 (Brian is now working with Dr. Michael Temkin)

Dan
DePuccio
: Woodburning and Indoor Air Quality (with Dr. Ning Gao)

Matthew
DePuccio with Connor Kellog
g: Effects of Liposomes Composed of
Phosphatidylcholine and Phosphatidyl-serine on the Structures of
Bovine Serum Albumin and Cytochrome C (with Dr. Nadia Marano)'

Michelle
Dumas
: BInding Specificity of Nogalamycin to Plasmid DNA (with Dr.
Samantha Glazier)

Josh
Johnson
: Lead Leaching in Ceramics and Glass: Impacts on Human
Health (with Dr. Ning Gao)

Ryan
Littlefield
: Synthesis of Novel Ligands for Copper 9-Metallacrown-3
Complexes (with Dr. Neil Law)

Chris
Lundberg
: Coordination Chemistry Investigations of Novel
Platinum(II)-Lanthanide(III) 15-Metallacrown-5 Complexes and
Prospective use as an MRI Contrasting Agent (with Dr. Neil Law)

Peterson
Maina
: Recruitment of RPD3 to the Promoter Sequences of Yeast Genes
(with Dr. Emily Dixon)
Geoff Millard: Design, Synthesis,
and Characterization of Metallacrown Compounds (with Dr. Neil Law)

Brian C. Palmer: Synthetic
Ruthenium Dimer Binding with DNA (with Dr. Samantha Glazier)

Alyssa
Pirinelli
: Boron Heterocycles: Boroles and Boronates (with Dr.
Larry French)

Hillary
St. John
: Biosensors and Androgen Receptors (with Dr. Dixon and Dr.
Skeels, Biochemistry)

Tim
Senter
: Synthesis of Novel N-Cyanoguanidine Vanillin Receptor
Ligands (with Dr. Larry French)

Marcus Tuttle: ITC for Analysis of
Ligand/DNA Complexes (with Dr. Samantha Glazier)

Cathy
Yancey
: Analysis of the Components of "5 Hour Energy" in a High
School Lab (with Dr. Larry French)

What function do both 14-3-3
proteins, Bmh1 and Bmh2, undertake in the transcriptional regulation of genes
in Saccharomyces cerevisiae during starvation in a nutrient limited
environment?

Samantha Berghorn with Dr. Emily Dixon, advisor

The yeast 14-3-3 proteins Bmh1 and Bmh2 alter the
transcription of genes under rapamycin treatment, an environment that mimics
nutrient-limited conditions in yeast. Nutrient-limited genes are regulated by
the TOR pathway in response to metabolic demands, nutrient availability, and
stress. Under nutrient-limited environments and rapamycin treatment, TOR
proteins are inhibited and transcription factors are localized to the nucleus,
which then up regulate nutrient-limited genes. Even though both of these 14-3-3
proteins overlap in function, a deletion of one but not necessarily both,
effects the growth and cell division of the yeast. A double deletion of both of
these proteins has been shown to be lethal in most but not all strains. Further
investigation reveals that both single deletions of Bmh1p and Bmh2p alter the
transcription of over 286 genes after treatment with rapamycin. A deletion of
one of these proteins will show the effect of that protein in absence of the
other but deleting both of these proteins will show their overall role during
nutrient-limited conditions. Since these proteins are shown to regulate
nutrient-limited genes in a single knockout, the principle behind this
experiment is to determine whether the double deletion of the 14-3-3 proteins
alters the transcription of the genes any differently then the single
deletions. Through a microarray analysis it has been determined that the strain
with the double knockout of both 14-3-3 proteins, BMH1 and BMH2, affects the
transcription of numerous genes throughout the entire genome, some different
and some the same as the single deletion strains.

Chemical Models of Anti-Cancer Pharmaceuticals
Based on Ruthenium Compounds
with Organic Side Chains. 

Kristin Berretta with Dr.
Samantha Glazier, advisor

Molecules that bind to DNA inhibit replication, which is important for
anti-cancer pharmaceuticals.  This work focuses on DNA binding analysis of
ruthenium compounds that have a modifiable chain attached to one of three
ligands surrounding the metal center.  Modifying the shape, chirality and
hydrophobicity of the binding molecule affects where binding occurs as well as
the strength of the interaction.  Generally molecules that bind strongly
and at specific locations are better candidates for drug-therapy.  The
syntheses of the modifiable ligand begins with 4-formyl-4’-methyl-bipyridine,
which is an aldehyde, produced using a selenium oxidation reaction with yields
up to 78%. The aldehyde is then reduced to an alcohol (95 % yield). The alcohol
undergoes a Williamson Ether synthesis to produce a bipyridine ligand with a
modifiable chain.  This ligand is bound to [Ru(phenanthroline)2Cl2] to
produce the final product shown in Figure 1.  All products were analyzed
by 1HNMR, 13CNMR, and infrared spectroscopy.  The [Ru(phen)2bpy-naphthalene](PF6)2
was bound to DNA in fluorescence titration studies.  The DNA binding
strengths were calculated and compared to a control ruthenium complex,
[Ru(phenanthroline)3](PF6)2.  The mode of binding is unknown and expected
to depend on the nature of the modified ligand.  We expect that binding
strength will increase with the addition of the naphthalene group compared to
control, because naphthalene is planar and aromatic that favors intercalation
between the base pairs of DNA. 

Figure 1. Final ruthenium compound, [Ru(phen)2(bpy-R)]2+. This Ru compound
has a charge of =2, which will attract the negatively charged DNA and a
modifiable bipyridine ligand expected to affect DNA binding mode and strength.

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Woodburning and Indoor Air
Quality.

Dan DePuccio with Dr. Ning Gao,
Advisor

In upstate New York, the burning of wood fuels for heating purposes is very
common in the winter months.  Woodstoves and pellet stoves are
conventional and practical options for residents seeking to heat their
homes.  Wood combustion may also pose a potential threat to indoor air
quality, and in effect, public health.  The purposes of this research are
threefold.  First, emissions of carbon monoxide, carbon dioxide, and
particulate matter (10 µm and 2.5 µm) are monitored at homes before (for
background) and during wood burning.  Second, the concentrations of toxic
species (such as formaldehyde and nitrogen oxides) released from each appliance
as a result from the combustion of wood are measured.  Polyaromatic
compounds in the emissions will be collected and analyzed through gas
chromatography-mass spectrometry.  Last, several wood and pellet samples
have been collected from the homes of volunteers and analyzed using adiabatic
oxygen bomb calorimetry to determine the gross heat of combustion for different
wood fuel types.  Analysis of all results will provide information on the
effects that wood burning has on air within residential buildings and the
heating efficiency of wood fuels.

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Effects of Liposomes Composed of Phosphatidylcholine
and Phosphatidyl-serine on the Structures of Bovine Serum Albumin and
Cytochrome C

Matthew DePuccio with Connor Kellogg
Advisor:  Dr. Nadia Marano

Abstract : Protein aggregation contributes to
neurodegenerative diseases such as Parkinson's and Alzheimer's disease. 
The earliest stages of prefibrillar aggregation are believed to have a
significant role in disease pathogenesis by increasing cytotoxicity within the
cell and disrupting physiological ion concentrations leading to eventual cell
death.  Phospholipids that make up cellular membranes are capable of
making proteins adopt non-native confirmations by causing the proteins to
initially unfold, and then allowing them to take up non-native conformations.  
Inferences can be made regarding the unfolding and folding processes of
proteins in the presence of phospholipids, specifically phosphatidylcholine
(PC) and phosphatidylserine (PS).  In order to study the effect of these
lipids on the structure of model proteins intrinsic fluorescence was used to
study changes in the environment surrounding the tryptophan residues within the
protein.  Fluorescence was measured when samples of bovine serum albumin
and cytochrome c were incubated in vitro with or without small-unilamellar
vesicles containing PS and PC for different times.  From initial studies,
PS had a larger affect on the structure of BSA and cytochrome c than PC and
mixed liposomes of PS/PC.  Next we will compare this observation with
changes in fluorescence brought about by urea, which denatures the
protein. 

Footprinting Assays to Study the
Binding Affinity of Nogalamycin to Various DNA Motifs.

Michelle Dumas with Dr. Samantha Glazier, advisor.

Cancer is
the uncontrolled proliferation of abnormal cells.  Highly conserved DNA
motifs are recognized by replication machinery in the promoter region of a gene
such as the stem cell leukemia gene (SCL) which act as regulatory elements
(Göttgens, 2002).  Through targeting these motifs and inhibiting DNA synthesis
of these regions with antibiotic drugs, cancer therapy could become more
selective.  Anthracycline drugs have been shown to treat various cancer
lines through intercalation.  Nogalamycin is an anthracycline antitumor
natural antibiotic that uses threading to inhibit DNA-directed RNA synthesis
which prevents a cell from dividing and proliferating.  The binding
mechanism of nogalamycin has been studied extensively, including SLU graduate
Matt Millard, but its specificity for various DNA structures has not been
determined.  Though the cardiotoxicity of nogalamycin will prevent its
commercial use, a similar compound could be synthesized with less adverse
effects.  Common techniques for studying DNA binding require large DNA
sample volumes.  Footprinting with topoisomerase and agarose gel
electrophoresis require much smaller samples but have only been used to study
intercalating molecules, never threading molecules (Webb, 2003).  Agarose
gel electrophoresis reveals the degree of intercalation post relaxation with
topoisomerase by the migration distance of the plasmid (Webb, 2003).  The
main focus of this research is to discover if this technique works with
threading molecules like nogalamycin and implementing it using a variety of DNA
motifs to determine the structural specificity of the antibiotic.

References:

Göttgens,
B., Barton, L.M., Chapman, M.A., Sinclair, A.M., Knudsen, B., Grafham, D.,
Gilbert, J.G.R., Rogers, J., Bentley, D., and Green, A. (2002). Transcriptional
Regulation of the Stem Cell Leukemia Gene (SCL): Comparative Analysis of
Five Vertebrate SCL Loci. Genome Res.12: 749-759.

Webb, M.R.
and Ebeler, S.E. (2003) A gel electrophoresis assay for the simultaneous
determination of topoisomerase I inhibition and DNA intercalation. Analy.
Biochem.
321: 22-30.

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How safe is your house?  An investigation of lead and cadmium levels in
household items.

Josh Johnson with Dr. Ning Gao, Advisor.

Lead and
cadmium, known as heavy metals, have been shown to have detrimental health
effects in humans, including bioaccumulation in many organs causing brain
damage, kidney damage, and gastrointestinal distress, as well as negative
reproductive and developmental effects.  The goal of this project is to
examine lead and cadmium levels in household items, primarily leaded crystal
glassware, glazed ceramics, and children’s toys, and how they might be ingested
through contact.  The project was created due to many product recalls in
recent years due to lead and cadmium levels.  The dinnerware items were
soaked in water, wine, brandy, tea and lemonade to imitate consumption and use
of the pieces.  The children’s toys were soaked in solutions of artificial
sweat and saliva to mimic toy-human contact by hand or by mouth. Analysis was
performed using the flame atomic absorption spectroscopy technique and the
results were compared to accepted values from the United States Consumer
Product Safety Commission and Food and Drug Administration.

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Synthesis of Novel Ligands
for Copper 9-Metallacrown-3 Complexes. 

Ryan Littlefield with Dr. Neil Law,
advisor

Metallacrowns are a unique family of inorganic molecules with metal-containing
ring structures at the center.  Oximes such as are one type of organic
molecule that can form metallacrowns, and this project explores the use of the
oxime phenyl 2- pyridyl ketoxime (pko).  This project began by
investigating the use of copper 9-metallacrown-3 (9-MC-3, 9 indicating the
total number of ring members with 3 metal atoms) complexes as dye sensitizers
for solar cells.  A researcher from another university suggested that
modifying the ligand by adding a carboxylic acid functional group on the phenyl
ring might improve the complex’s utility as a dye sensitizer.  These
molecules, however, did not show positive results, and modifying the ligand
would not have helped.  However, no carboxylate derivatives of pko have
been utilized to form metallacrowns.  This project’s overall goal is now
the synthesis of modified pko ligands for use in the synthesis of an inverse
copper 9-MC-3 complex.   Another interesting feature of this project
would be that the modified pko ligand could allow the metallacrowns to be
linked together by coordinating the carboxylic acids to other metal ions. 
This could then produce extended solids or molecular containers.

At least three synthetic routes are possible for the new ligand.  Two
of the more common have been tried.  The first multistep synthetic route
was adapted from a procedure reported in 1998 by Takeuchi , et al., that
couples 3-bromopyridine with methyl 4-formylbenzoate.   Product was
not achieved in sighnificant amounts when 3-bromopyridine was replaced with 2-bromopyridine
to create the modified phpyko.  When tested, the 3-bromopyridine synthesis
worked as well as had been reported.  The current approach was outlined by
Villani and Papa.   This route uses a Friedel-Crafts acylation followed
by oxidation with potassium permanganate.   Results from these
synthetic approaches and any new metallacrowns will be presented.

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Coordination Chemistry Investigations of Novel Platinum(II)-Lanthanide(III)
15-Metallacrown-5 Complexes and Prospective use as an MRI Contrasting Agent
.

Christopher J. Lundberg
with Dr. Neil Law, Advisor

Hydroxamic acids coordinated to first row transition metals is
well documented in the chemical literature, specifically in relation to
copper(II), iron(III), and nickel(II), but there is very little concerning
hydroxamic coordination with soft less labile second and third row transition
metals such as platinum and palladium. The first goal of this study explores
the synthesis and characterization of five previously cited platinum hydroxamic
acid complexes using 195Pt-NMR; an aptitude of nuclear magnetic resonance
spectroscopy that has never been collected on this class of molecules. The
prior compounds show (O,O), (O,N), and (N,N) hydroxamate coordination modes
about the platinum and were discovered to coordinate to platinum via
deprotonated hydroxyl and carbonyl oxygen atoms giving rise to stable five
member chelate rings. The second goal of this study explores numerous coordination
chemistry investigations of platinum in an attempt to synthesize the first
platinum metallacrown; a molecule that has not been previously reported in the
chemical literature. Metallacrowns are unique symmetric inorganic compounds
with ring structures at their core. Metallacrowns based on hydroxamic acids
form a cyclic ring structure consisting of an alternating metal ion (e.g. Pt),
oxygen atom (O), and nitrogen atom (N) respectively (Pt-O-N), with a metal ion
or lanthanide ion (La) encapsulated at its core. Metallacrowns are presently
being investigated for use as chiral recognition agents, catalysts,
antibiotics, single molecular magnets, and MRI contrasting agents. Water
molecules in the human body bind to the lanthanide ion in commercially used
contrasting agents such as [Gd(DOTA)]2-, which enhances the relaxation of water
protons and is measured by the MRI machine and used to create an image. In
lanthanide-metallacrowns, up to four water molecules can bind to the central
lanthanide ion compared to one water molecule in commercially used contrasting
agents. When more water molecules are able to bind to the lanthanide, a clearer
and more defined MRI image is possible. Currently no metallacrowns are used as
MRI contrasting agents because metallacrowns tend to degrade and become labile
inside the pH of the human body. Platinum is a harder less labile metal
compared to first row transition metals currently used to synthesize
metallacrowns. Platinum has been shown to remain intact and withstand the thermodynamic
requirements of human tissue in that it positively intercalates DNA and is
currently used in 50% of all anti-cancer treatments.

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What Transcription Factor is
sufficiently necessary for binding of HDAC Rpd3 onto the
PAC Promoter Element in Yeast?

Peterson Maina with Dr. Emily Dixon,
advisor

Rapamycin
treatment of Saccharomyces cerevisiae (yeast) cells has been shown to
regulate genes involved in nutrition pathways. Such regulation often involves
activation or repression of certain proteins, and it is barely understood.
Herein, we investigated the recruitment of the Histone Deacetylase (HDAC)
enzyme RPD3 onto the promoter sequences of nutrition associated genes,
hypothesizing that certain transcription factors linked to RPD3 recruit it to
these specific promoter sequences. Computational analysis of these promoter
sequences revealed a common CTCATC motif (the Polymerase A and C element- PAC
element) shown to bind PBF1 and PBF2 transcription factors. Reverse
Transcription Quantitative PCR of GAP1, RPA43 and RPC25 genes on doubly deleted
PBF1 and PBF2 cells compared to Wild Type cells showed transcription profiles
slightly similar to those of RPD3.

To
demonstrate sufficiency of the PAC element in recruiting, a strain of cells
without GAP1 gene was unsuccessfully developed via transformation using DNA
pringle cassette. Mutagenesis of the same GAP1 gene on a plasmid was
inconclusive too. However, microarray experiment on the doubly deleted PBF1 and
PBF2 transcription factors revealed the regulation of ribosomal protein genes.
This analysis in essence increases understanding of a mechanism that could be
mimicked in cancerous growth.

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Boron Containing Heterocycles: Suitability of Dioxaboroles and Boronate
Esters
in Diels-Alder Reactions.

Alyssa Pirinelli with Dr.
French, Advisor

Boron containing compounds are useful in synthesis reactions
because of their versatility and affinity for Lewis acid base chemistry and are
found in natural as well as in complex synthetic products.  Usually found
in a trigonal planar bonding geometry, boron has an empty p-orbital
perpendicular to the bonding plane.  In the presence of a Lewis base,
boron will adopt a tetrahedral geometry and a negative charge and can for
instance form a boronate ester.  The p-orbital can also promote
aromaticity by allowing circulation of electron density around the ring, as is
the case for a dioxaborole.  Both boronate esters and dioxaboroles are
heterocyclic ring systems, with boron attached to two alkyl-bridged oxygen
atoms.  The dioxaborole system is aromatic in nature, with electron pair
donation from the two oxygen atoms and an alkene pi bond, using the boron to
share electron density.  Boronate esters have alkane chains connecting the
two oxygen atoms, however, and are therefore non-aromatic.  Instead, the
boronate esters are useful in coordination chemistry, allowing reagents to
tether to one another through boron, causing it to switch bonding geometry. 
Both of these heterocycles types are useful in Diels-Alder reactions, though
they perform opposite roles.  Dioxaboroles are used as dienophiles while
boronate esters act as dienes, the function dependent on what the boron is
attached to.  In boronate ester chemistry, because a dienophile such as
methyl vinyl ketone coordinates with boron through a lone pair on the ketone,
an intramolecular Diels-Alder reaction may then happen between the alkene bond
and the furan.  Attempts at synthesis of various boroles and boronate
esters have so far been unsuccessful.

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The Development of a New Biosensor for Detecting Androgen Receptor
Agonists and Antagonists Based on FRET Analysis.

Hillary St. John with advisors
Drs. Emily Dixon, Biology and Matthew Skeels, Chemistry.

Abstract:  The androgen receptor is a transcription factor with three
domains.  The carboxy-terminal domain is the androgen-binding domain,
which is activated by the binding of one of several select steroid hormones,
such as testosterone.  The activation of the androgen receptor by the
hormone ligand interaction causes the protein to induce the transcription of
androgen mediated genes.  My project focuses on the conformational changes
of the androgen-binding domain upon ligand binding.  We have a construct
consisting of the androgen-binding domain of the androgen receptor linked with
fluorophores on both the N-terminus and C-terminus of the domain.  The
goal of the project is to obtain the construct in high yield and subsequently
use FRET (fluorescence resonance energy transfer) techniques to investigate the
conformational changes of the protein in the presence of various synthetic
steroid hormone-like and natural steroid hormone ligands. 

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Synthesis of Novel N-cyanoguanidine
Analogs of Transient Receptor Potential Vanillin Type-1 antagonists: Impact of
Incorporation of D-Region Binding Sites

Timothy Senter, with Dr. Larry French, advisor

Abstract:
This project consists of the synthesis and analysis of novel molecules that
feature a newly proposed binding site for interaction with the Transient
Receptor Potential Vanillin Type-1 (TRPV1) Receptor. The human TRPV1 receptor
is responsible for transmitting the sensation of pain when triggered by noxious
stimuli. These stimuli include capsaicin (the active ingredient in chili
peppers), endovanilloids (chemicals released during tissue inflammation and
some diseases that cause pain signals to be sent to the brain), low pH and heat
(ii). The proposed molecules have been designed to effectively bind to this
receptor, blocking the interaction between the noxious stimuli and the TRPV1
receptor without causing a response from the receptor. Molecules that behave in
this manner are referred to as antagonists (iii). Through this mechanism, the
novel molecules work as analgesics to relieve pain in people who suffer from
chronic pain conditions such as osteoarthritis, multiple sclerosis, diabetes,
and complications due to cancer. A number of such TRPV1 antagonists have
progressed into human clinical studies. A synthetic route, based on the
utilization of dimethyldithiocyano-iminocarbonate to faciltate linkage of two
large aminosubstituted precursors via a cyanoguanidine group will be used to
create these molecules. These antagonists feature the proposed D-region binding
site (iv) that is expected to increase affinity to the TRPV1 receptor. After
the molecules have been synthesized and purified, their activities will be
measured in a nematode bioassay. Results will be compared to previously
synthesized antagonists from Dr. French's lab to determine the effectiveness of
the D-region binding site. This research will help to build upon recent
discoveries in the field of analgesic research and serve as a template for
further exploration of TRPV1 antagonists containing the D-region binding site.

i. Szallasi,
Arpad. (2003) Journal of Medicinal Chemistry, (47) 11, 2717-2723
ii. Szallasi, Arpad, et al. (2007) Nature, (6) 5, 357-372
iii. Tamayo, Nurai, et al. (2008) Journal of Medicinal Chemistry (51),
2744-2757
iv. Lee, Jeewoo. (2001) Bioorganic and Medicinal Chemistry (9) 19-32

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Analysis of the Components of
"5 Hour Energy" in a High School Lab

Cathy Yancy with Dr. French, advisor

Abstract:
Caffeine is a very common and well known compound.  It is the perfect
compound for high school students to study because of the prior knowledge they
already possess.  In the experiment that is being developed, students will
be able to extract caffeine from 5 Hour Energy (a popular energy drink) using
liquid extraction.  They will then recrystallize the caffeine and verify
the identity using TLC.  TLC will also be used to verify the presence of
the other ingredients of 5 Hour Energy including amino acids, vitamins, and
sugars.  Lessons will be developed that allow the students to study the
biochemical aspects of each of the ingredients.  Students will be able to
decide if energy drinks are good for them, based on the information they gain.

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