title: Cucurbit[7]uril-Tetramethylrhodamine Conjugate for Direct Sensing and Cellular Imaging.
abstract: This paper describes the design and synthesis of a conjugate (Q7R) comprising the synthetic host cucurbit[7]uril (Q7) linked to the fluorescent dye tetramethylrhodamine (TMR), and the characterization of its optical and guest-binding properties as well as its cellular uptake. Q7R was synthesized in two steps from monofunctionalized azidobutyl-Q7 and NHS-activated TMR. The fluorescence of Q7R is quenched upon guest binding, and this observable was used to determine equilibrium dissociation constant (Kd) values. Unexpectedly, the Kdvalues for guests binding to Q7R and to unmodified Q7 were essentially identical. Therefore, Q7R can directly report binding to Q7 without an energetic penalty due to the conjugated fluorophore. This result demonstrates a potentially general strategy for the design of single-component host-indicator conjugates that respond sensitively to analytes without perturbing the binding properties of the host. The unique properties of Q7R enabled measurement of Kdvalues across 3 orders of magnitude and at concentrations as low as 0.7 nM. This result is particularly relevant given the unmatched range of guests and binding affinities demonstrated for Q7. Confocal fluorescence microscopy of live and fixed HT22 neurons revealed the cellular uptake of Q7R and its punctate localization in the cytoplasm. Q7R did not alter cell growth at concentrations up to 2.2 μM over 4 days. These experiments demonstrate the feasibility of Q7R as a direct sensor for guest binding and as a cell-permeable compound for imaging applications.
authors.forename: Andrew T
authors.initials: AT
authors.lastname: Bockus
authors.forename: Lauren C
authors.identifier: 0000-0002-4290-3262
authors.initials: LC
authors.lastname: Smith
authors.forename: Amy G
authors.initials: AG
authors.lastname: Grice
authors.forename: Omar A
authors.initials: OA
authors.lastname: Ali
authors.forename: Carolyn C
authors.initials: CC
authors.lastname: Young
authors.forename: William
authors.initials: W
authors.lastname: Mobley
authors.forename: Ashley
authors.initials: A
authors.lastname: Leek
authors.forename: James L
authors.initials: JL
authors.lastname: Roberts
authors.affiliation: Department of Chemistry and Biochemistry, University of Maryland , College Park, Maryland 20742, United States.
authors.forename: Brittany
authors.initials: B
authors.lastname: Vinciguerra
authors.affiliation: Department of Chemistry and Biochemistry, University of Maryland , College Park, Maryland 20742, United States.
authors.forename: Lyle
authors.identifier: 0000-0002-4079-332X
authors.initials: L
authors.lastname: Isaacs
authors.forename: Adam R
authors.identifier: 0000-0003-3426-4760
authors.initials: AR
authors.lastname: Urbach
mesh_terms: D001952:Bridged-Ring Compounds / Q000737:chemistry*; D045744:Cell Line, Tumor; D006801:Humans; D007093:Imidazoles / Q000737:chemistry*; D057054:Molecular Imaging / Q000379:methods*; D012235:Rhodamines / Q000737:chemistry*; D047028:Macrocyclic Compounds; D000099246:Cucurbit(n)urils
chemical_list: D001952:Bridged-Ring Compounds; D007093:Imidazoles; D012235:Rhodamines; C456276:cucurbit(7)uril; D047028:Macrocyclic Compounds; D000099246:Cucurbit(n)urils; C005358:tetramethylrhodamine