Category Archives: Seminars

Assessing the Binding Capabilities of Bromodomain-Containing Protein 9

Sarah Hopson (Advisor- Dr. Martin Thompson)

Doctoral Student, Department of Chemistry,Michigan Technological University

Monday, March 2, 2015-9:00 am- Admin 404

Abstract

Post-translational modifications of histones, such as the acetylation of lysines, play an importantrole in regulating transcription. Histone tails have a large proportion of positively-charged residues, which create electrostatic interactions with the negatively-charged DNA backbone. Lysine acetylation is thought to weaken these interactions, because it neutralizes lysine’s positively-charged side chain.

Proteins recognize the acetylated lysines using bromodomains; bromodomains are acetylated lysine “readers” and play a critical role in modulation of gene expression. Of the 46 bromodomain-containing proteins in the human proteome, 15 function as transcriptional regulators and 8 function as chromatin remodelers. Nearly all of the other bromodomain proteins influence transcription in some manner (histone acetyltransferase, transcription repressor, transcription initiation, etc.). Due to their significant influence on transcription, mutations of bromodomains are often linked with cancers.

Bromodomain-containing protein 9 (BRD9) has not yet been studied. The aim of this proposed research is to determine the specificity and affinity of BRD9 toward acetyl-lysine sites on the tails of the four core histone proteins.

A high-throughput examination of possible histone interactions with the bromodomain of BRD9 will be conducted using a modified SPOT array. The peptides demonstrating the strongest interactions with the bromodomain will be synthesized using standard Fmoc peptide synthesis. A quantitative examination of the binding affinities of these peptides to the bromodomain, the bromodomain and DUF3512 (domain of unknown function), and the full length BRD9 will be conducted using isothermal titration calorimetry. The results will be compared to determine how the surrounding amino acid sequences affect the bromodomain’s binding capabilities.


Bio-Oligomer Purification, Electrophilic Oligo Synthesis, and Progress to a Genome Sequencer

CHEMISTRY SEMINAR

Michigan Technological University

Friday, December 12, 2014

3:00 pm Chemical Sciences Building Room 101

Dr. Shiyue Fang

Associate Professor, Chemistry Department

Abstract:

Bio-Oligomer Purification, Electrophilic Oligo Synthesis, and Progress to a Genome Sequencer

The progress on three projects, which are bio-oligomer purification, electrophilic oligosynthesis, and developing a new genome sequencer, will be presented. For bio-oligomer purification, we have developed two methods for oligodeoxynucleotide and one method for peptide purification. They are catching failure sequences by polymerization and catching full-length sequences by polymerization. Both methods do not require any type of chromatography,and purification is achieved through simple manipulations such as shaking and filtration. As aresult, they are suitable for large scale purification of drugs based on oligonucleotides and peptides. They are also ideal for small scale purification and high throughput purification. Currently, there are three oligonucleotide drugs and over 60 peptide drugs on the market, and many more are in various stages of clinical trials. Because known bio-oligomer purification methods such as HPLC have drawbacks such as high capital cost for instrument, labor-intensiveness and requirement of large volumes of harmful solvents, the new methods are expected to be preferred by pharmaceutical companies for drug purification, and by academic labs and biotech companies for small scale and high throughput purification. For electrophilic oligo synthesis, we have made progress on developing a new method that features using protecting groups and linkers cleavable under nearly neutral conditions. Under these conditions, electrophilic groups such as ester, thioester, alpha-halo carbonyl, epoxide and aziridine are stable. As a result, the new method is useful for the synthesis of oligonucleotide analogs that contain such  sensitive functionalities. Using known oligo synthesis methods, such analogs cannot be synthesized. The new oligo synthesis method is expected to open doors to many research projects that require  sensitive oligo analogs. For developing a new genome sequencer, we are using AFM to monitor the  conformational fluctuations of a DNA polymerase during DNA synthesis. Because different  nucleotides are expected to give different conformational fluctuations, DNA sequences can be read  out in real-time. We have made progress on mutating a polymerase and solving several potential  problems for assembling the sequencer.


NON-CHROMATOGRAPHIC PURIFICATION OF SYNTHETIC BIOOLIGOMERS

Durga Pokharel

Advisor: Dr. Shiyue Fang

Doctoral candidate, Department of Chemistry

PhD Defense

Friday December 12, 2014   9:30am     Fisher 130

 

NON-CHROMATOGRAPHIC PURIFICATION OF SYNTHETIC BIOOLIGOMERS

Abstract

 

Synthetic oligonucleotides and peptides have found wide applications in industry and academic research labs. There are ~60 peptide drugs on the market and over 500 under development. The global annual sale of peptide drugs in 2010 was estimated to be $13 billion. There are three oligonucleotide-based drugs on market; among them, the FDA newly approved Kynamro was predicted to have a $100 million annual sale. The annual sale of oligonucleotides to academic labs was estimated to be $700 million. Both bio-oligomers are mostly synthesized on automated synthesizers using solid phase synthesis technology, in which nucleoside or amino acid monomers are added sequentially until the desired full-length sequence is reached. The additions cannot be complete, which generates truncated undesired failure sequences. For almost all applications, these impurities must be removed. The most widely used method is HPLC. However, the method is slow, expensive, labor-intensive, not amendable for automation, difficult to scale up, and unsuitable for high throughput purification. It needs large capital investment, and consumes large volumes of harmful solvents. The purification costs are estimated to be more than 50% of total production costs. Other methods for bio-oligomer purification also have drawbacks, and are less favored than HPLC for most applications.

To overcome the problems of known biopolymer purification technologies, we have developed two non-chromatographic purification methods. They are (1) catching failure sequences by polymerization, and (2) catching full-length sequences by polymerization. In the first method, a polymerizable group is attached to the failure sequences of the bio-oligomers during automated synthesis; purification is achieved by simply polymerizing the failure sequences into an insoluble gel and extracting full-length sequences. In the second method, a polymerizable group is attached to the full-length sequences, which are then incorporated into a polymer; impurities are removed by washing, and pure product is cleaved from polymer. These methods do not need chromatography, and all drawbacks of HPLC no longer exist. Using them, purification is achieved by simple manipulations such as shaking and extraction. Therefore, they are suitable for large scale purification of oligonucleotide and peptide drugs, and also ideal for high throughput purification, which currently has a high demand for research projects involving total gene synthesis. The savings with the new techniques compared with HPLC are estimated to be 70% to 90% depending on purification scale and throughput. We expect these new oligonucleotide and peptide purification technologies to be widely used in academic research labs, biotechnology companies, and pharmaceutical companies in the near future.


Purification and Carbohydrate Binding Properties of Two New Plant Proteins

Mr. Robert K Brown

Advisor: Dr. Tarun K Dam

Master’s Candidate Department of Chemistry

Michigan Technological University

“Purification and Carbohydrate Binding Properties of Two New Plant Proteins”

 

Friday, December 12, 2014

10:00 – 11:00 AM 

Room 404 ~ Administration Building
Purification and Carbohydrate Binding Properties of Two New Plant Proteins

Abstract:

 

Protein glycosylation is an important post-translational modification for many biological processes such as cell recognition, intercellular communication and cell death. Proteins that are able to bind to glycosylated proteins via carbohydrates are called lectins. Hemolytic lectins are proteins or glycoproteins that undergo specific interactions with cell surface carbohydrates and subsequently induce cellular lysis. They are termed “hemo” lytic because of their ability to lyse erythrocytes. We have isolated a novel hemolytic lectin named HelyX from the bulbs of a monocot plant, as well as a mannose-binding lectin named DIL from a separate monocot species. HelyX is a uniquely robust hemolytic lectin. It shows concentration dependent reversible hemolytic/agglutinating properties against both human and rabbit erythrocytes. The activity was found to be carbohydrate dependent. HelyX was isolated using ammonium sulfate precipitation, size exclusion chromatography, and analyzed by gel electrophoresis. DIL was purified using a modified version of a newly developed protocol. DIL interacts with the plant enzyme invertase with high affinity. This high affinity interaction suggests that the binding site of DIL is complimentary to glycoproteins containing larger high mannose glycans. Invertase is central to plant metabolism and defense. Therefore DIL might play a modulatory role in plant metabolism and defense through its interaction with invertase. HelyX and DIL did not show lytic activity on free living amoeba, Acanthamoebae. Instead the lectins promoted cyst formation of amoeabae trophozoites indicating a lectin-mediated rearrangement of membrane architecture. This result indicates that the lytic activity of HelyX or DIL depends on the macromolecular landscape of the cell membrane.


Evolution of Selected Isoprene Oxidation Products in Dark Aqueous Ammonium Sulfate

MS Defense:  DM Ashraf Ul Habib
Chemistry Department

Lynn Mazzoleni, Advisor
Thursday, December 4  1pm,  Chem Sci 101
Evolution of Selected Isoprene Oxidation Products in Dark Aqueous Ammonium Sulfate

The climate of the world is changing but our understanding of atmospheric processes is limited. Atmospheric aerosol is a trace but very influential medium of the atmosphere. Especially little is known about the organic aerosol components and their aqueous phase chemical evolution. To address this, the aqueous phase processing of glyoxylic acid, py­ruvic acid, oxalic acid and methylglyoxal was studied simulating dark and radical free atmospheric aqueous aerosol. A novel observation of the cleavage of a carbon-carbon bond in pyruvic acid and glyoxylic acid leading to their decarboxylation was made in the presence of ammonium salts but decarboxylation was not observed from oxalic acid. The empirical rate constants for decarboxylation were determined and are competitive with nighttime OH radical reactions. The structure of the acid, ionic environment of the solu­tions and concentration of species were all found to affect the rate of decarboxylation. A tentative set of reaction mechanisms is proposed involving nucleophilic attack by ammo­nia on the carbonyl carbon leading to fragmentation of the carbon-carbon bond between the carbonyl and carboxyl carbons. Under similar conditions in atmospheric aerosol, the aqueous phase processing may markedly impact the physicochemical properties of aerosol.

 


Synthetic Oligodeoxynucleotide Purification via Catching by Polymerization

Suntara (Boat) Fueangfung

Advisor: Dr. Shiyue Fang

Doctoral candidate, Department of Chemistry

Michigan Technological University

Friday, December 5, 2014, 9:00 am

Admin Building, Room 404

PhD Defense

 

Synthetic Oligodeoxynucleotide Purification via Catching by Polymerization

Abstract

Large quantities of pure synthetic oligodeoxynucleotides (ODNs) are important for preclinical research, drug development, and biological studies. These ODNs are synthesized on an automated synthesizer. It is inevitable that the crude ODN product contains failure sequences which are not easily removed because they have the same properties as the full length ODNs. Current ODN purification methods such as polyacrylamide gel electrophoresis (PAGE), reversed-phase high performance liquid chromatography (RP HPLC), anion exchange HPLC, and affinity purification can remove those impurities. However, they are not suitable for large scale purification due to the expensive aspects associated with instrumentation, solvent demand, and high labor costs.

To solve these problems, two non-chromatographic ODN purification methods have been developed. In the first method, the full-length ODN was tagged with the phosphoramidite containing a methacrylamide group and a cleavable linker while the failure sequences were not. The full-length ODN was incorporated into a polymer through radical acrylamide polymerization whereas failure sequences and other impurities were removed by washing. Pure full-length ODN was obtained by cleaving it from the polymer. In the second method, the failure sequences were capped by a methacrylated phosphoramidite in each synthetic cycle. During purification, the failure sequences were separated from the full-length ODN by radical acrylamide polymerization. The full-length ODN was obtained via water extraction. For both methods, excellent purification yields were achieved and the purity of ODNs was very satisfactory. Thus, this new technology is expected to be beneficial for large scale ODN purification.


Lake Superior’s History and Future

 

On Tuesday, Nov. 18, Professor Sarah Green, expert on Lake Superior, will lead a discussion titled “Lake Superior’s History and Future,” at the Carnegie Museum. Refreshments will be served at 6:30 p.m., with the discussion following at 7 p.m.

The event is part of a monthly series of sessions on the Geoheritage and Natural History of the Keweenaw, at the Carnegie Museum in Houghton. The discussions are aimed at the general public, but discuss current research and science.


Title: Applications of Quantum Chemical Methods to Atmospheric Reactions

Dr. Heather A. Rypkema

Department of Atmospheric, Oceanic, and Space Sciences

University of Michigan

 

November 14, 2014  

3:00 pm Chemical Sciences Building~ Room 101

Abstract:

Quantum chemical methods have a broad-ranging capacity toward informing our understanding of molecular transformations in a variety of environments, including the chemistry of the atmosphere. Specifically, these applications include kinetic and thermodynamic analysis of chemical reactions, evaluation of competing reaction channels, and the theoretical validation of postulated chemical mechanisms. This presentation provides a survey quantum chemistry as applied to a number of atmospherically relevant studies. The full atmospheric oxidation profile of peroxyacetic acid predicts the likely fate of a prevalent atmospheric species while postulating a new source of reactive species not represented in current models. The direct and catalyzed hydration of formic acid and acetaldehyde provide a mechanism for the formation of hygroscopic molecules capable of initiating cloud formation and the production of SOA. Diabatic excited states can be used to predict the relative reactivity among atom-transfer reactions, which are particularly significant in a hydroxyl-rich environment. A mechanism for the proliferation of atmospheric hydroxyl radical through the atmospheric oxidation of cyclical alkanes provides a possible explanation for the discrepancy between predicted and measured OH levels in the troposphere. Cumulatively, these studies will demonstrate the diverse applications of theoretical quantum chemistry in enhancing our understanding of atmospheric chemistry.

 

 

 


Elucidating the Pathways for Protein Misfolding and Aggregation: Unity in Diversity.

Dr. Ashutosh Tiwari, Assistant Professor of Chemistry

Department of Chemistry,  Michigan Technological University     

 

November 7, 2014  ~  3:00pm  ~  Chemical Science Building, Room 101

Abstract:

Due to the aging of baby boomers in the USA, the proportion of the population in higher age groups has increased. This demographic shift coupled with a concomitant increase in longevity has brought new challenges and threats in the form of diseases and disorders that not only affect an individual but impact the whole society at large. Increased oxidative damage of proteins associated with aging causes them to misfold and aggregate and thus, disorders related to protein misfolding and aggregation are on the rise. Since many aggregated proteins share a common fibrillar structure at the molecular level, understanding the principles and contributing factors that regulate protein misfolding, surface hydrophobic exposure, aberrant interactions, or aggregation is key to understanding their relationship to cellular toxicity. I will discuss recent results from my laboratory wherein we studied several proteins for their surface-hydrophobic exposure and aggregation propensity at physiological pH and temperature. Identifying shared protein aggregation pathways for a large set of structurally diverse proteins will lead to a better understanding of the disease process and as a consequence provide common effective targets for therapy.

 

 


Green Chemistry: An Overview of Principles and Applications

Mark R. Mason

Professor, Department of Chemistry and Biochemistry and
Director,  School of Green Chemistry and Engineering
The University of Toledo
Thursday, November 6, 2014
11:00 am Chemical Sciences Building Room 102

Over the past two decades, there has been a dramatic shift in the way government and industry view pollution prevention and the environmental consequences of chemical manufacture in the United States. Source reduction, rather than “end of the pipe” waste treatment, is now the preferred method for reducing pollution. This approach requires chemists and engineers to be increasingly aware of the environmental consequences of the chemical-related products and processes we develop. Green chemistry, “the design of chemical products and processes that reduce or eliminate the use and generation of hazardous substances,” is the foundation of this increased awareness. This presentation will provide an overview of selected green chemistry principles, green chemistry applications and metrics, chemical alternatives assessment, and future opportunities.