Basic Amino Acids as Charged State Reducing Agents for Native Mass Spectrometry
Disciplines
Biochemistry
Abstract (300 words maximum)
Native Mass Spectrometry (MS) allows for the characterization of biomolecules in their native state, preserving their structure. The electrospray ionization of the native mass spectrometry transfers samples from the liquid phase to the gas phase by introducing a high voltage to the sample. The electrostatic charge of the sample causes proteins to unfold, leading to a more complex mixture of ions, complicating data analysis, identification efforts, and quantificational efforts. To prevent the unfolding of proteins charged state reducers can be used to lower the electrostatic repulsion between highly charged groups on the protein which maximizes the maintenance of the native conformation of proteins. In previous studies, it has been found that high concentrations of imidazole derivatives are effective in lowering the charge state that is acquired during the ionization process resulting in lowered repulsion between charged groups of a protein as well as dissociation. In this research, the charge reducing capacity of various basic amino acids such as histidine, lysine, and arginine are tested for a model protein lysozyme. The mass spectrum of lysozyme obtained in water showed the charge states from 8+ to 11+ where 10+ charge state was more dominant. Various concentrations of amino acids were tested from 0.1 mM to 1 mM. In the presence of Histidine, Lysozyme showed the charge states from 4+ to 10+ where 7+ was more intense. Similar features were obtained for Arginine and Lysine. This study showed that basic amino acids have potential to be very effective charge reducing agents and particularly significant charge reducing capacity was noticed for Histidine compared to all other amino acids and imidazole.
Academic department under which the project should be listed
CSM - Chemistry and Biochemistry
Primary Investigator (PI) Name
Mohammad A. Halim
Basic Amino Acids as Charged State Reducing Agents for Native Mass Spectrometry
Native Mass Spectrometry (MS) allows for the characterization of biomolecules in their native state, preserving their structure. The electrospray ionization of the native mass spectrometry transfers samples from the liquid phase to the gas phase by introducing a high voltage to the sample. The electrostatic charge of the sample causes proteins to unfold, leading to a more complex mixture of ions, complicating data analysis, identification efforts, and quantificational efforts. To prevent the unfolding of proteins charged state reducers can be used to lower the electrostatic repulsion between highly charged groups on the protein which maximizes the maintenance of the native conformation of proteins. In previous studies, it has been found that high concentrations of imidazole derivatives are effective in lowering the charge state that is acquired during the ionization process resulting in lowered repulsion between charged groups of a protein as well as dissociation. In this research, the charge reducing capacity of various basic amino acids such as histidine, lysine, and arginine are tested for a model protein lysozyme. The mass spectrum of lysozyme obtained in water showed the charge states from 8+ to 11+ where 10+ charge state was more dominant. Various concentrations of amino acids were tested from 0.1 mM to 1 mM. In the presence of Histidine, Lysozyme showed the charge states from 4+ to 10+ where 7+ was more intense. Similar features were obtained for Arginine and Lysine. This study showed that basic amino acids have potential to be very effective charge reducing agents and particularly significant charge reducing capacity was noticed for Histidine compared to all other amino acids and imidazole.
