A method for generating targeted, pattern-generating, protein surface sensors via the self-assembly of modified oligodeoxynucleotides (ODNs) is described. The simplicity by which these systems can be created enabled the development of a sensor that can straightforwardly discriminate between distinct glycoform populations. By using this sensor to identify glycosylation states of a therapeutic protein, we demonstrate the diagnostic potential of this approach as well as the feasibility of integrating a wealth of supramolecular receptors and sensors into higher-order molecular analytical devices with advanced properties. For example, the facile device integration was used to attach the well-known anthracene-boronic acid (An-BA) probe to a biomimetic DNA scaffold and consequently, to use the unique photophysical properties of An-BA to improve glycoform differentiation. In addition, the noncovalent assembly enabled us to modify the sensor with a trinitrilotriacetic acid (tri-NTA)-Ni2+ complex, which endows it with selectivity toward a hexa-histidine tag (His-tag). The selective responses of the system to diverse His-tag-labeled proteins further demonstrate the potential applicability of such sensors and validate the mechanism underlying their function.
Anthracenes/chemistry , Boronic Acids/chemistry , Oligodeoxyribonucleotides/chemistry , Periplasmic Binding Proteins/analysis , Glycosylation , Molecular Structure , Oligodeoxyribonucleotides/chemical synthesis , Surface Properties
Fluorescent molecular probes have become valuable tools in protein research; however, the current methods for using these probes are less suitable for analysing specific populations of proteins in their native environment. In this study, we address this gap by developing a unimolecular fluorescent probe that combines the properties of small-molecule-based probes and cross-reactive sensor arrays (the so-called chemical 'noses/tongues'). On the one hand, the probe can detect different proteins by generating unique identification (ID) patterns, akin to cross-reactive arrays. On the other hand, its unimolecular scaffold and selective binding enable this ID-generating probe to identify combinations of specific protein families within complex mixtures and to discriminate among isoforms in living cells, where macroscopic arrays cannot access. The ability to recycle the molecular device and use it to track several binding interactions simultaneously further demonstrates how this approach could expand the fluorescent toolbox currently used to detect and image proteins.
Electronic Nose , Fluorescent Dyes/chemistry , Proteins/analysis , Proteins/chemistry
Signal transduction pathways, which control the response of cells to various environmental signals, are mediated by the function of signaling proteins that interact with each other and activate one other with high specificity. Synthetic agents that mimic the function of these proteins might therefore be used to generate unnatural signal transduction steps and consequently, alter the cell's function. We present guidelines for designing 'chemical transducers' that can induce artificial communication between native proteins. In addition, we present detailed protocols for synthesizing and testing a specific 'transducer', which can induce communication between two unrelated proteins: platelet-derived growth-factor (PDGF) and glutathione-S-transferase (GST). The way by which this unnatural PDGF-GST communication could be used to control the cleavage of an anticancer prodrug is also presented, indicating the potential for using such systems in 'artificial signal transduction therapy'. This work is intended to facilitate developing additional 'transducers' of this class, which may be used to mediate intracellular protein-protein communication and consequently, to induce artificial cell signaling pathways.
Antineoplastic Agents/metabolism , Aptamers, Nucleotide/pharmacology , Azo Compounds/metabolism , Chemistry Techniques, Synthetic , Glutathione Transferase/metabolism , Piperazines/metabolism , Platelet-Derived Growth Factor/metabolism , Prodrugs , Signal Transduction/drug effects , Aptamers, Nucleotide/chemical synthesis , Cell Physiological Phenomena , Glutathione Transferase/chemistry , Nitric Oxide/metabolism , Platelet-Derived Growth Factor/chemistry , Prodrugs/metabolism
Antineoplastic Agents/pharmacology , Neoplasms/drug therapy , Protein Kinase Inhibitors/pharmacology , Receptor Protein-Tyrosine Kinases/antagonists & inhibitors , Signal Transduction/drug effects , Antineoplastic Agents/chemistry , Humans , Neoplasms/metabolism , Neoplasms/pathology , Protein Kinase Inhibitors/chemistry , Receptor Protein-Tyrosine Kinases/metabolism
The design and function of a synthetic "chemical transducer" that can generate an unnatural communication channel between two proteins is described. Specifically, we show how this transducer enables platelet-derived growth factor to trigger (in vitro) the catalytic activity of glutathione-s-transferase (GST), which is not its natural enzyme partner. GST activity can be further controlled by adding specific oligonucleotides that switch the enzymatic reaction on and off. We also demonstrate that a molecular machine, which can regulate the function of an enzyme, could be used to change the way a prodrug is activated in a "programmable" manner.
Glutathione Transferase/antagonists & inhibitors , Oligonucleotides/pharmacology , Platelet-Derived Growth Factor/metabolism , Cell Line, Tumor , Cell Survival/drug effects , Dose-Response Relationship, Drug , Enzyme Activation/drug effects , Glutathione Transferase/metabolism , Humans , Models, Molecular , Molecular Structure , Oligonucleotides/chemistry , Oligonucleotides/metabolism , Platelet-Derived Growth Factor/chemistry , Protein Binding/drug effects , Structure-Activity Relationship
