| Abstract
| - We have designed rhenium(I) biotin isothiocyanate complexes as the first class of luminescent biotinylation reagents. The photophysical properties of these complexes and their amine and thiourea counterparts have been investigated. Bovine serum albumin has been biotinylated with the isothiocyanate complexes, and the photophysical properties of the resultant bioconjugates have been studied. The cytotoxicity of the thiourea complexes towards HeLa cells and the cellular uptake of one of the thiourea complexes have also been examined.
- We report here the design of the first class of luminescent biotinylation reagents derived from rhenium(I) polypyridinecomplexes. These complexes [Re(N−N)(CO)3(py-biotin-NCS)](PF6) (py-biotin-NCS = 3-isothiocyanato-5-(N-((2-biotinamido)ethyl)aminocarbonyl)pyridine; N−N = 1,10-phenanthroline (phen) (1a), 3,4,7,8-tetramethyl-1,10-phenanthroline (Me4-phen) (2a), 4,7-diphenyl-1,10-phenanthroline (Ph2-phen) (3a)), containing a biotin unit and anisothiocyanate moiety, have been synthesized from the precursor amine complexes [Re(N−N)(CO)3(py-biotin-NH2)](PF6) (py-biotin-NH2 = 3-amino-5-(N-((2-biotinamido)ethyl)aminocarbonyl)pyridine; N−N = phen (1c), Me4-phen(2c), Ph2-phen (3c)). To investigate the amine-specific reactivity of the isothiocyanate complexes 1a−3a, they havebeen reacted with a model substrate ethylamine, resulting in the formation of the thiourea complexes [Re(N−N)(CO)3(py-biotin-TU-Et)](PF6) (py-biotin-TU-Et = 3-ethylthioureidyl-5-(N-((2-biotinamido)ethyl)aminocarbonyl)pyridine;N−N = phen (1b), Me4-phen (2b), Ph2-phen (3b)). All the rhenium(I) complexes have been characterized, andtheir photophysical properties have been studied. The avidin-binding properties of the thiourea complexes 1b−3bhave been examined by the 4‘-hydroxyazobenzene-2-carboxylic acid (HABA) assay. Titration results indicated thatthe complexes exhibited emission enhancement by ca. 1.4−1.5-fold upon binding to avidin, and the lifetimes wereelongated to ca. 0.8−2.0 μs. Additionally, we have biotinylated bovine serum albumin (BSA) with the isothiocyanatecomplexes. All the resultant rhenium−BSA bioconjugates displayed intense and long-lived orange-yellow to greenish-yellow emission upon irradiation in aqueous buffer under ambient conditions. The avidin-binding properties of thebioconjugates have been investigated using the HABA assay. Furthermore, the cytotoxicity of the thiourea complexes1b−3b toward the HeLa cells has been examined by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide(MTT) assay. The IC50 values were determined to be ca. 17.5−28.5 μM, which are comparable to that of cisplatin(26.7 μM) under the same conditions. The cellular uptake of complex 3b has been investigated by fluorescencemicroscopy, and the results showed that the complex was localized in the perinuclear region after interiorization.
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