Cell Permeable Peptides

21st Century Biochemicals Scientists Are Co-Authors on Two Cell-Permeable Peptide Papers! A recent paper in the J. Biological Chemistry from the laboratory of John Johnson at the Medical College of GA demonstrated the ability of cell-permeable peptides derived from PKC delta to modify the activity of the F1F0-ATPase in mitochondria. This study clearly demonstrates the ability to deliver peptide cargo to the mitochondria and to either activate or inhibit enzyme activity. The close collaboration between Dr. Johnson's laboratory and Dr. Jordan Fishman is indicative of the collaborative relationship between the scientists at 21st Century Biochemicals and our customers around the world. See: Modulation of the Delta Protein Kinase C Interaction With The "d" Subunit of F1Fo ATP Synthase in Neonatal Cardiac Myocytes: Development of Cell-Permeable, Mitochondrial-Targeted Inhibitor and Facilitator Peptides. Tiffany T. Nguyen, Mourad Ogbi, Qilin Yu, Jordan B. Fishman, Warren Thomas, David Fulton, and John A. Johnson. July, 2010. J. Biol. Chem., 285: 22164 - 22173

In a second paper, the laboratory of Dr. Peter Polgar, along with Drs. Berg, Shia and Fishman of 21st Century Biochemicals demonstrate that peptides can be used to modulate angiotensin receptor signaling through the calcium/PI turnover while having no effect of ERK activation. The use of cell-permeable peptides to delineate regulation of various arms of receptor signaling offers an advantage over the use of siRNA and similar knock-down technologies, where multiple signaling pathways would be affected. See: Limiting angiotensin II signaling with a cell penetrating peptide mimicking the second intracellular loop of the angiotensin II type I receptor. Jun Yu, Linda Taylor, Dale Mierke, Eric Berg, Michael Shia, Jordan Fishman, and Peter Polgar. 2010. Chem. Biol. & Drug Design, Chem Biol Drug Des, July 1, 2010; 76(1): 70-6.

Cell Permeable Peptides - An Introduction

Intracellular signal transduction is mediated in large part by the interaction(s) and subsequent modification(s), e.g., phosphorylation, of key regulatory proteins. Protein complexes resulting from the juxtaposition of anywhere from just two to as many as a dozen or more proteins control a myriad of cellular functions. In order to further study the interaction(s) of these proteins and their role in regulating cellular function, the use of cell permeable peptides has been employed as a means to introduce into the cell small pieces of these proteins (i.e., peptides) in order to disrupt the interaction of these proteins and study the downstream events.

Small basic (i.e., lysine and/or arginine-rich) peptides derived from the transduction domains of a number of proteins, including the third transmembrane domain of the Antennapedia homeodomain (also known as Penatratin) and the HIV-tat protein, can cross cell membranes without the need for a receptor or other transport carrier. Other mechanisms to introduce peptides into cells include using polyarginine peptides as mimics of the HIV-tat protein, the addition of a fatty acid, such as palmitic or stearic acid or the addition of other hydrophobic groups. The addition of one or more of the above vectors allows certain molecules to become cell-permeable In addition to modifying peptides, other biologically active molecules such as DNA or proteins can be introduced into cells.

The ultimate destination for this material can be determined through the use of general cell-permeability vectors, such as those above, or through the creation of directed vectors, which might contain for example nuclear localization signals, resulting in the transport of the material of interest to the nucleus. It is always important to keep in mind that design of these molecules requires careful consideration, as not all molecules will in fact be able to enter cells or will be delivered to the intended cellular compartment. It is advisable in some cases to add dyes or other molecules to confirm that the material has been introduced into the cell. In addition, it might be necessary to utilize more than one delivery strategy as well as the use of a scrambled control to rule out nonspecific effects due to the presence of the various components of the system being used.

In cases where the proteins that are interacting is known but the exact site(s) at which the proteins interact is unknown, protein-protein interactions can be studied through the use of zero-length, self-hydrolyzing crosslinkers. In general, a series of overlapping, lower purity peptides can be used first in a broken/permeablized cell system to determine the site(s) of interaction using crosslinkers are introduced to covalently link proteins that are in close proximity. Antibodies or other methods to recover the protein-peptide complex (i.e., biotinylated peptide, epitope-tagged protein) can then be employed to immunoprecipitate protein complexes of interest, and the MS lab at 21st Century Biochemicals can map the site(s) of interaction between the proteins via proteolytic fragment analysis. Once the sites have been identified, suitable cell-permeable peptides can be made to try and disrupt the association between the proteins of interest.

The design of cell-permeable molecules can be achieved in a cost-efficient manner if the projects initial stages are well thought out. The scientists at 21st Century Biochemicals have extensive experience in the design and production of peptides modified with cell-permeabilizing vectors, and can provide expert technical assistance. Together with our experienced mass spectrometry lab and peptide synthesis and purification labs, we are able to provide one stop shopping for your protemics needs.




 

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