Translational and rotational motions of albumin sensed by a non-covalent associated porphyrin under physiological and acidic conditions: a fluorescence correlation spectroscopy and time resolved anisotropy study.
The interaction between a free-base, anionic water-soluble porphyrin, TSPP, and the drug carrier protein, bovine serum albumin (BSA) has been studied by time-resolved fluorescence anisotropy (TRFA) and fluorescence correlation spectroscopy (FCS) at two different pH-values. Both rotational correlation times and translational diffusion times of the fluorescent species indicate that TSPP binding to albumin induces very little conformational changes in the protein under physiological conditions. By contrast, at low pH, a bi-exponential decay is obtained where a short rotational correlation time (phi (int) = 1.2 ns) is obtained, which is likely associated to wobbling movement of the porphyrin in the protein binding site. These physical changes are corroborated by circular dichroism (CD) data which show a 37% loss in the protein helicity upon acidification of the medium. In the presence of excess porphyrin formation of porphyrin J-aggregates is induced, which can be detected by time-resolved fluorescence with short characteristic times. This is also reflected in FCS data by an increase in molecular brightness together with a decrease in the number of fluorescent molecules passing through the detection volume of the sample.
Andrade
Suzana M
SM
Centro de Química Estrutural, Complexo 1, Instituto Superior Técnico, Lisbon, Portugal. suzana.andrade@ist.utl.pt
Costa
Silvia M B
SM
Borst
Jan Willem
JW
van Hoek
Arie
A
Visser
Antonie J W G
AJ
D000465:Algorithms; D001665:Binding Sites; D002942:Circular Dichroism; D005454:Fluorescence Polarization; D006863:Hydrogen-Ion Concentration; D011166:Porphyrins / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D012710:Serum Albumin, Bovine / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence
D011166:Porphyrins; D012710:Serum Albumin, Bovine; C016136:tetraphenylporphine sulfonate