Conformation and dynamics of bovine brain S-100a protein determined by fluorescence spectroscopy.
We have used time-resolved laser fluorescence spectroscopy to investigate the intensity and anisotropy decays of the single tryptophan residue in bovine brain S-100a (alpha beta) protein. The steady-state and acrylamide quenching results indicated that the Trp 90 of the alpha-subunit was partially buried in a relatively nonpolar environment at pH 7.5. Both Ca2+ and pH 8.5 slightly enhanced the exposure of the residue to the solvent, but the residue remained partially buried in the calcium complex at both pH values. The best representation of the intensity decays was a linear combination of three exponential terms, regardless of solvent condition and temperature. The three lifetimes (tau i) were in the range of 0.4-5 ns and insensitive to emission wavelength, but their fractional amplitudes (alpha i) shifted in favor of the shortest component (alpha 1) when the decays were measured at the blue end of the emission spectrum. These results suggest that an excited-state interaction between the indole ring and the side chain of an adjacent residue may be responsible for the observed shortest lifetime. In the presence of Ca2+, the three lifetimes remained relatively unaltered, but the values of alpha 1 decreased by a factor of 2.3 at pH 7.2 and a factor of 1.8 at pH 8.2. This Ca(2+)-induced decrease may be attributed to disruption of the putative excited-state interaction resulting from reorientations of the alpha-helical segments flanking a Ca(2+)-binding loop (residues 62-73). At both pH 7.2 and 8.4, the anisotropy decays of the apoprotein followed a biexponential decay law.(ABSTRACT TRUNCATED AT 250 WORDS)
Wang
C K
CK
Department of Physiology and Biophysics, University of Washington, Seattle 98195.
Mani
R S
RS
Kay
C M
CM
Cheung
H C
HC
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D005454:Fluorescence Polarization; D006863:Hydrogen-Ion Concentration; D009038:Motion; D011487:Protein Conformation; D009418:S100 Proteins / Q000737:chemistry / Q000648:ultrastructure*; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000737:chemistry
D009418:S100 Proteins; D014364:Tryptophan; D002118:Calcium