ja0469737_si_001.pdf (506.88 kB)
Perturbation of Fluorescence by Nonspecific Interactions between Anionic Poly(phenylenevinylene)s and Proteins: Implications for Biosensors
journal contributionposted on 2004-12-29, 00:00 authored by Stephen J. Dwight, Brent S. Gaylord, Janice W. Hong, Guillermo C. Bazan
The use of anionic water-soluble conjugated polymers (CPs) for sensing the presence of avidin by use of a biotin-modified fluorescence quencher was studied. The molecules involved in the study included poly[2-methoxy-5-(3‘-propyloxysulfonate)-1,4-phenylenevinylene] with either lithium (Li+-MPS-PPV) or sodium (Na+-MPS-PPV) countercations, the well-defined oligomer pentasodium 1,4-bis(4‘(2‘ ‘,4‘ ‘-bis(butoxysulfonate)-styryl)-styryl)2-butoxysulfonate-5-methoxybenzene (5R5-), the quenchers N-methyl-4,4‘-pyridylpyridinium iodide (mMV+) and [N-(biotinoyl)-N‘-(acetyl 4,4‘-pyridylpyridinium iodide)] ethylenediamine (BPP+), which contains a molecular recognition fragment (biotin) attached to a unit that accepts an electron from a CP excited state, and the proteins avidin, tau, BSA, and pepsin A. Fluorescence quenching experiments were examined in a variety of conditions. Experiments carried out in water and in ammonium carbonate buffer (which ensures avidin/biotin complexation) reveal that nonspecific interactions between the CP and the proteins cause substantial perturbations on the CP fluorescence. The overall findings are not consistent with a simple mechanism whereby avidin complexation of BPP+ leads to encapsulation of the quencher molecule and recovery of Li+-MPS-PPV fluorescence. Instead, we propose that binding of BPP+ to avidin results in the quenching unit attaching to a positively charged macromolecule. Electrostatic attraction to the negatively charged conjugated polymer results in closer proximity to the quencher. Therefore, more enhanced fluorescence quenching is observed.
quencher moleculerecognition fragmentammonium carbonate bufferiodideavidin resultsfluorescence quenchingproteins causeavidin complexationquenchers NMPSPPV fluorescenceElectrostatic attractionCP fluorescenceNonspecific InteractionsBSAproteins avidinLiBPPFluorescence quenching experimentspolymer resultsquenching unit5 R 5