Synthesis and Characterization, and Label-Free Quantification of Cell Penetrating Peptide in Yeast Cell
Disciplines
Analytical Chemistry | Biochemistry
Abstract (300 words maximum)
Cell-penetrating peptides (CPPs) are short sequences of amino acids (5-30) possessing a positive charge, enabling their entry into cells by passing through the cell membrane and being high in amino acids like arginine and lysine. Those factors present a huge potential for biomedical applications including transportation of cargo into cells via endocytosis. CPPs have been employed as carriers for delivering proteins or genes into various cells and tissues. They are good at penetrating their membrane walls, but it can be complex to do this without damaging or destroying the cell. In this course-based research project, three short peptide sequences of CPPs were chosen to be synthesized, characterized by mass spectrometry, and performed label free liquid chromatography and mass spectrometry assay to quantify how many cell-perpetrating peptides enter inside the Yeast cell. These peptides were synthesized using a CEM Liberty Blue peptide synthesizer, peptide-resin drying and cleavage using 95% trifluoracetic acids, and the peptides being filtered and precipitated with cold diethyl ether. Peptide characterization was then conducted with mass spectrometry. Electrospray ionization-mass spectrometry (ESI-MS) is an analytical method that is used to determine the molecular weight of these peptides. The mass spectrometry results can confirm the successful synthesis of each of the peptides as the expected charge states were seen using mass spectrometer. CPP1 that was synthesized showed one strong peak at m/z 864 which corresponds to the [M+H]+ charge state. CPP2 demonstrated a dominant peak at m/z 670 which corresponds to the [M+H]+ charge state. Likewise, a strong peak at m/z 763 was noticed for CPP3 which agreed with the theoretical masses. Currently, label free LCMS assay is under investigation to quantify the CPP entrance to the Yeast cell.
Academic department under which the project should be listed
CSM - Chemistry and Biochemistry
Primary Investigator (PI) Name
Mohammad A. Halim
Synthesis and Characterization, and Label-Free Quantification of Cell Penetrating Peptide in Yeast Cell
Cell-penetrating peptides (CPPs) are short sequences of amino acids (5-30) possessing a positive charge, enabling their entry into cells by passing through the cell membrane and being high in amino acids like arginine and lysine. Those factors present a huge potential for biomedical applications including transportation of cargo into cells via endocytosis. CPPs have been employed as carriers for delivering proteins or genes into various cells and tissues. They are good at penetrating their membrane walls, but it can be complex to do this without damaging or destroying the cell. In this course-based research project, three short peptide sequences of CPPs were chosen to be synthesized, characterized by mass spectrometry, and performed label free liquid chromatography and mass spectrometry assay to quantify how many cell-perpetrating peptides enter inside the Yeast cell. These peptides were synthesized using a CEM Liberty Blue peptide synthesizer, peptide-resin drying and cleavage using 95% trifluoracetic acids, and the peptides being filtered and precipitated with cold diethyl ether. Peptide characterization was then conducted with mass spectrometry. Electrospray ionization-mass spectrometry (ESI-MS) is an analytical method that is used to determine the molecular weight of these peptides. The mass spectrometry results can confirm the successful synthesis of each of the peptides as the expected charge states were seen using mass spectrometer. CPP1 that was synthesized showed one strong peak at m/z 864 which corresponds to the [M+H]+ charge state. CPP2 demonstrated a dominant peak at m/z 670 which corresponds to the [M+H]+ charge state. Likewise, a strong peak at m/z 763 was noticed for CPP3 which agreed with the theoretical masses. Currently, label free LCMS assay is under investigation to quantify the CPP entrance to the Yeast cell.