Chemically reactive derivatives of gramicidin A for developing ion channel-based nanoprobes.
Ion channel-forming peptides and proteins offer tremendous opportunities for fundamental and applied studies of function on individual molecules. An ongoing challenge in ion channel research is the lack of simple and accessible synthetic methods to engineer pores with tailored chemical and physical properties. This paper describes a practical synthetic route to rapidly generate C-terminally modified derivatives of gramicidin A (gA), an ion channel-forming peptide, through the use of two chemically reactive gA-based building blocks. These amine- and azide-containing building blocks can react readily with typical substrates for amidation and 1,3-dipolar cycloaddition ("click") reactions to present molecules with desired structure and functionality near the opening of a gA pore. These derivatives of gA are stable under typical aqueous conditions for ion channel recordings and retain characteristic single ion channel conductance properties in planar lipid bilayers. Additionally, the synthetic methods described here afford useful quantities of these gA derivatives in good purity and yield with minimal purification. We demonstrate that derivatives of gA can be used for studying, in situ, a change in conductance through a channel upon performing a "click" reaction on an azide moiety attached to the gA pore. We also demonstrate that these gA-based building blocks can be used to construct sensors for the recognition of specific protein-ligand binding interactions in solution. This widely accessible, enabling synthetic methodology represents a powerful new tool to study relationships between chemical structure and function on the single molecule level.
Blake
Steven
S
Department of Chemistry and Biochemistry, University of California, San Diego, 9500 Gilman Drive, MC 0358, La Jolla, California 92093-0358, USA.
Capone
Ricardo
R
Mayer
Michael
M
Yang
Jerry
J
D000595:Amino Acid Sequence; D015374:Biosensing Techniques / Q000379:methods*; D006096:Gramicidin / Q000031:analogs & derivatives / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D007473:Ion Channels / Q000378:metabolism*; D008024:Ligands; D008969:Molecular Sequence Data; D049329:Nanostructures / Q000737:chemistry*; D011485:Protein Binding
D007473:Ion Channels; D008024:Ligands; D006096:Gramicidin