Investigating the Impact of Glucose, Sucrose and Glycerol on Protein Structure and Dynamics using Electrospray Ionization-Mass Spectrometry
Disciplines
Analytical Chemistry | Biochemistry
Abstract (300 words maximum)
Protein, antibody, peptide and enzyme-based medications remarkably increased in the pharmaceutical industry, but long-term storage and stability issues arise with newer and more complex medications. In the last two decades, the FDA approved 894 therapeutic proteins, including 354 monoclonal antibodies and 85 peptides or polypeptides. A critical aspect of this is the preservation of these protein-based therapeutics, which require various stabilizers that can maintain their stability. Various stabilizers such as sucrose, glucose and glycerol are frequently used to storge and preserve these medications. The goal of this research is to investigate how these stabilizing agents’ impact on the folding and unfolding of a model protein. Various concentrations of these stabilizing agents were prepared and mixed with Lysozyme and incubated and analyzed by electrospray ionization coupled with mass spectrometry techniques. Unfolded lysozyme, which is less stable, leads to substantially higher charge states than the folded one. All mass spectrums were obtained using an LTQ XL mass spectrometer. In this experiment, the control sample, Lysozyme in water, exhibited m/z peaks ranging from 8+ to 11+, with the strongest peak at 10+. When Lysozyme was incubated with various concentrations (1 mM to 50 mM) of glucose, Lysozyme exhibited m/z peaks ranging from 8+ to 13+, with the strongest peak at 12+ which indicates that the protein is unfolded in presence of glucose. Similar features were noticed when Lysozyme was incubated with sucrose, however, various adduct peaks emerged which confirm that sucrose is not only unfolding the protein but also glycosylating the lysozyme significantly. While comparing the results with glucose and sucrose, this is evident that glycerol showed m/z peaks ranging from 8+ to 12+ with the strongest peak at 10+ which showed that glycerol was the most effective on stabilizing the protein.
Academic department under which the project should be listed
CSM - Chemistry and Biochemistry
Primary Investigator (PI) Name
Mohammad A. Halim
Investigating the Impact of Glucose, Sucrose and Glycerol on Protein Structure and Dynamics using Electrospray Ionization-Mass Spectrometry
Protein, antibody, peptide and enzyme-based medications remarkably increased in the pharmaceutical industry, but long-term storage and stability issues arise with newer and more complex medications. In the last two decades, the FDA approved 894 therapeutic proteins, including 354 monoclonal antibodies and 85 peptides or polypeptides. A critical aspect of this is the preservation of these protein-based therapeutics, which require various stabilizers that can maintain their stability. Various stabilizers such as sucrose, glucose and glycerol are frequently used to storge and preserve these medications. The goal of this research is to investigate how these stabilizing agents’ impact on the folding and unfolding of a model protein. Various concentrations of these stabilizing agents were prepared and mixed with Lysozyme and incubated and analyzed by electrospray ionization coupled with mass spectrometry techniques. Unfolded lysozyme, which is less stable, leads to substantially higher charge states than the folded one. All mass spectrums were obtained using an LTQ XL mass spectrometer. In this experiment, the control sample, Lysozyme in water, exhibited m/z peaks ranging from 8+ to 11+, with the strongest peak at 10+. When Lysozyme was incubated with various concentrations (1 mM to 50 mM) of glucose, Lysozyme exhibited m/z peaks ranging from 8+ to 13+, with the strongest peak at 12+ which indicates that the protein is unfolded in presence of glucose. Similar features were noticed when Lysozyme was incubated with sucrose, however, various adduct peaks emerged which confirm that sucrose is not only unfolding the protein but also glycosylating the lysozyme significantly. While comparing the results with glucose and sucrose, this is evident that glycerol showed m/z peaks ranging from 8+ to 12+ with the strongest peak at 10+ which showed that glycerol was the most effective on stabilizing the protein.