Native serotonin membrane receptors recognize 5-hydroxytryptophan-functionalized substrates: enabling small-molecule recognition.
Recognition of small diffusible molecules by large biomolecules is ubiquitous in biology. To investigate these interactions, it is important to be able to immobilize small ligands on substrates; however, preserving recognition by biomolecule-binding partners under these circumstances is challenging. We have developed methods to modify substrates with serotonin, a small-molecule neurotransmitter important in brain function and psychiatric disorders. To mimic soluble serotonin, we attached its amino acid precursor, 5-hydroxytryptophan, via the ancillary carboxyl group to oligo(ethylene glycol)-terminated alkanethiols self-assembled on gold. Anti-5-hydroxytryptophan antibodies recognize these substrates, demonstrating bioavailability. Interestingly, 5-hydroxytryptophan-functionalized surfaces capture membrane-associated serotonin receptors enantiospecifically. By contrast, surfaces functionalized with serotonin itself fail to bind serotonin receptors. We infer that recognition by biomolecules evolved to distinguish small-molecule ligands in solution requires tethering of the latter via ectopic moieties. Membrane proteins, which are notoriously difficult to isolate, or other binding partners can be captured for identification, mapping, expression, and other purposes using this generalizable approach.
Department of Bioengineering Huck Institutes of the Life Sciences, Pennsylvania State University, University Park, Pennsylvania 16802, USA.
Amit
A
Vaish
Mitchell J
MJ
Shuster
Sarawut
S
Cheunkar
Yogesh S
YS
Singh
Paul S
PS
Weiss
Anne M
AM
Andrews
D006916:5-Hydroxytryptophan / Q000276:immunology / Q000378:metabolism*; D000327:Adsorption; D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000378:metabolism; D000888:Antibodies, Anti-Idiotypic / Q000378:metabolism; D000918:Antibody Specificity; D001682:Biological Availability; D002417:Cattle; D005026:Ethylene Glycols; D006046:Gold; D007074:Immunoglobulin G / Q000378:metabolism; D008024:Ligands; D008958:Models, Molecular; D011485:Protein Binding; D011817:Rabbits; D011985:Receptors, Serotonin / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000378:metabolism; D012996:Solutions; D013379:Substrate Specificity; D013438:Sulfhydryl Compounds; D013499:Surface Properties
D000906:Antibodies; D000888:Antibodies, Anti-Idiotypic; D005026:Ethylene Glycols; D007074:Immunoglobulin G; D008024:Ligands; D011985:Receptors, Serotonin; D012996:Solutions; D013438:Sulfhydryl Compounds; C077067:hexaethylene glycol; D012710:Serum Albumin, Bovine; D006046:Gold; D006916:5-Hydroxytryptophan