Time-resolved fluorescence of the single tryptophan residue in rat alpha-fetoprotein and rat serum albumin: analysis by the maximum-entropy method.
The time-resolved fluorescence emissions of the lone tryptophan residues in rat alpha-fetoprotein (RFP) and rat serum albumin (RSA) were studied. The total fluorescence intensity decays in both proteins were multiexponential. Analysis of the data by nonlinear least squares as a sum of discrete exponentials showed that four exponentials were needed for a satisfactory fit for both proteins. Analysis by the maximum entropy method using 150 logarithmically equally spaced exponentials yielded four well-resolved excited-state lifetime classes with barycenters and relative amplitudes values (ci) that corresponded to those obtained from the nonlinear least-squares method. Changing the temperature affected the relative amplitudes of the lifetime classes but had little effect on the lifetime values themselves. This suggests that the four classes reflect local conformational substates that exchange slowly with respect to the time window of observation defined by the longest lifetime. The internal rotational dynamics of the tryptophan in each protein was monitored by fluorescence anisotropy decay measurements. The mobility of the tryptophan appeared to be larger and faster in RFP than in RSA. The nonlinear least-squares analysis suggests the existence of three rotational correlation times of 0.1, 3, and 55 ns for this protein. As a function of temperature, the long correlation time did not follow the Perrin's law expected for a rigid rotating body. This suggests that this correlation time may reflect not only the Brownian rotation of the whole protein but also the flexibilities of domains in the protein. For RSA a two-component model with correlation times of 0.4 and 31 ns was sufficient to describe the data.(ABSTRACT TRUNCATED AT 250 WORDS)
Gentin
M
M
Laboratoire pour l'Utilisation du Rayonnement Electromagnétique, Centre National de la Recherche Scientifique, Université Paris-Sud, Orsay, France.
Vincent
M
M
Brochon
J C
JC
Livesey
A K
AK
Cittanova
N
N
Gallay
J
J
D000818:Animals; D004583:Electrons; D005454:Fluorescence Polarization; D051381:Rats; D012709:Serum Albumin / Q000737:chemistry*; D013816:Thermodynamics; D014364:Tryptophan / Q000737:chemistry*; D000509:alpha-Fetoproteins / Q000737:chemistry*
D012709:Serum Albumin; D000509:alpha-Fetoproteins; D014364:Tryptophan