Jessica Daniel*, Aviva Bulow, Susan Jett*, Jonathan Richards*, Laura Roon*, Kathryn Norquest*, Stephen Schaffner*, Ryan Masterson*, Ilia Mazin, Nazar Dubchak, Ryan Warren, Tonya Santaus, Kyra Brandt, Elina Baravik*, Josh Sowick, Becky Addison, Jody Stephens*, Yerelsy Reyna*, Jeremy O’Brien, Travis Ingraham, Matthew Stoddard, Morgan Miller, Amanda Faux, Mason Preusser, Sarai Graves, Tiffany Ashbaugh, Michael McCoy, Ebony Miller
(* denotes a researcher with a publication from the lab)
Dr. Andrew J. Bonham
Associate Professor of Chemistry
Dr. Bonham’s Curriculum Vitae
Dr. Bonham’s work focuses on understanding and investigating Transcription Factors, essential human proteins that regulate the bodies growth and response to disease. These transcription factors are essential components of gene regulation, and there is great interest in probing their presence and activity in both academic analysis and clinical diagnostics. Current methods to address these questions are often time-intensive or require specialized reagents, such as antibodies. At Metropolitan State University of Denver, Dr. Bonham is leading an innovative undergraduate research program focused on engineering new tools for sensitive and quick detection of TF:DNA interactions.
Lab Member 2014-
Current Project: Selecting aptamers for the detection of the cancer biomarker ENOX2
Former Project: Expanding electrochemical DNA biosensors to detect ricin
Ricin toxin chain A (RTA), a byproduct of the production of castor oil from castor bean plants, is a hazardous toxin that inhibits the cellular production of proteins once it enters the body. This toxin, whether ingested, inhaled, or injected, can be lethal, and treatment is difficult as there are currently no known antidotes for ricin. Detection of RTA prior to exposure is thus important, and it is necessary to expand the methods that can be used for this detection, as current methods are not time-sensitive. In this project, we have designed and tested an electrochemical biosensor that is capable of– and sensitive enough– to detect small, bio-medically relevant concentrations of RTA. This biosensor was designed based on an existing oligonucleotide aptamers that have been previously shown to bind to the hydrolase protein in ricin toxin. One of these aptamers was then used as the basis of a rationally designed DNA oligonucleotide biosensor scaffold that allows the coupling of RTA binding to a conformational change in the oligonucleotide. Ultimately, this biosensor design allows voltammetric interrogation to detect RTA concentration in complex media (such as coffee, blood, and river water). Additionally, this biosensor is convenient and collects real-time data, offering beneficial applications to the monitoring processes of areas involved in castor oil production. Furthermore, it may possess diagnostic potential in assessing ricin exposure. Electrochemical DNA biosensors have the potential to be used in numerous different situations, and this project shows a strategy for how they may be expanded to the detection and quantification of hazardous toxins.
Mycoplasma pneumonia infects 2 million people every year and is responsible for upper respiratory infections and “walking pneumonia.” Here, we describe the creation of a novel electrochemical biosensor capable of detecting pathogenic Mycoplasma for use in academic, research, and clinical applications. Current diagnostics of Mycoplasma, such as molecular-based assays, PCR and serological analysis, are time consuming, expensive, and not particularly accurate. In response, our biosensor is designed for rapid, reliable, and reagentless detection of several common Mycoplasma strains. To do so, we rely on the fact that many pathogenic Mycoplasma share a common secreted protein, P48. A modified aptamer against P48 was incorporated into a custom oligonucleotide scaffold and is used in a gold-electrode-bound fashion to give electrochemical signal change upon binding the secreted P48 target. Ultimately, this biosensor should bring improvements to diagnosis and thus treatment of Mycoplasma in patients who present a proposed infection.
One of the many great challenges that medical diagnostics face is the need for sensitive, reliable, and rapid detection of molecules in very complex solutions such as blood or urine. DNA-based biosensors have shown great promise in terms of sensitivity and reliability for target detection, but the need for rapid testing has considerably slowed their use in practical applications within the medical world. In the research to be conducted, we explore the incorporation of DNA-based biosensors into a lateral flow assay format (similar to the common at-home pregnancy test for human chorionic gonadotropin in urine). To facilitate this, we are developing a gold nanoparticle decorated with a functional DNA probe that recognizes and binds to botulism neurotoxin variant A (BoNTA). This conjugate then wicks across a nitrocellulose membrane to specific capture points, allowing rapid visual assessment of the BoNTA contamination of a sample. In the future, we aim to demonstrate that this represents a generic platform for detection that could be used with any existing DNA aptamer-based biosensing technique and can be applied to many medical settings, including small clinics, without the need for technicians to operate the biosensor.
Celiac disease is an autoimmune disorder where ingesting dietary gluten causes an autoimmune response that leads to villous atrophy and other complications. Symptoms can have gastrointestinal or extraintestinal manifestations. The symptoms can include: diarrhea, abdominal pain, dental enamel defects, neurological issues, Dermatitis Herpetiformis and more. This disease affects an estimated 2 million Americans, 1.4 million of which are undiagnosed1. Rate of diagnosis is low because many affected individuals have “silent celiac disease” where the symptoms are not detectable, but villous atrophy is still occurring. Also, there is a broad spectrum of symptoms for the condition and is often misdiagnosed. The average time it takes to diagnosis individuals who are symptomatic in the US is 11 years2. In that time, their risk of developing other autoimmune disorders, neurological problems, fertility problems and other conditions increases. One key challenge of this condition is that the diagnosis has historically suffered from difficulties in performing non-invasive positive identification of affected patients. Recently, interacting amino acid sequences on the surface of human tissue transglutaminase and gliadin proteins from foodstuffs have been identified as the potential recognition sites of commonly developed auto-antibodies in Celiac disease. In this project, we are exploring the use of short synthetic peptides that mimic these key amino acid recognition sites in order to create rapid, non-invasive, electrochemical biosensors to aid in the early diagnosis of Celiac disease. These biosensors build on a literature of electrochemical biosensors that offer reagentless, reusable, and rapid testing directly in small quantities of blood (such as a finger lancet draw). Our goal is to design, synthesize, and validate such diagnostic biosensors.
Gold nanoparticles, typically particles with a diameter of < 100 nanometers, present unique electrical and optical properties from the bulk material. These properties have been used to enable a variety of sophisticated molecular and chemical techniques. One of the most intriguing of these is the phenomenon of surface-enhanced Raman scattering, wherein closely spaced nanoparticles dramatically increase the sensitivity of Raman, a technique that allows “fingerprinting” of molecules. In this project, we synthesized gold nanoparticles, coated them with single stranded thiolated DNA. In the near future, we will anneal two variations of functionalized gold nanoparticles together. A functionalized Gold nanoparticle scaffold will form and promote signal Raman amplification due to calculated spacing. Designated proteins will bind to specific regions of DNA in the scaffold. binding should alter the molecular fingerprint of an organic dye, methylene blue, that is attached to the DNA. Ultimately, such assemblies should offer a stable optical platform for the creation of incredibly sensitive biosensors.