Monomer-dimer equilibrium in glutathione transferases: a critical re-examination.
Glutathione transferases (GSTs) are dimeric enzymes involved in cell detoxification versus many endogenous toxic compounds and xenobiotics. In addition, single monomers of GSTs appear to be involved in particular protein-protein interactions as in the case of the pi class GST that regulates the apoptotic process by means of a GST-c-Jun N-terminal kinase complex. Thus, the dimer-monomer transition of GSTs may have important physiological relevance, but many studies reached contrasting conclusions both about the modality and extension of this event and about the catalytic competence of a single subunit. This paper re-examines the monomer-dimer question in light of novel experiments and old observations. Recent papers claimed the existence of a predominant monomeric and active species among pi, alpha, and mu class GSTs at 20-40 nM dilution levels, reporting dissociation constants (K(d)) for dimeric GST of 5.1, 0.34, and 0.16 microM, respectively. However, we demonstrate here that only traces of monomers could be found at these concentrations since all these enzymes display K(d) values of <<1 nM, values thousands of times lower than those reported previously. Time-resolved and steady-state fluorescence anisotropy experiments, two-photon fluorescence correlation spectroscopy, kinetic studies, and docking simulations have been used to reach such conclusions. Our results also indicate that there is no clear evidence of the existence of a fully active monomer. Conversely, many data strongly support the idea that the monomeric form is scarcely active or fully inactive.
Department of Chemical Sciences and Technologies, University of Rome Tor Vergata, 00133 Rome, Italy.
Raffaele
R
Fabrini
Anastasia
A
De Luca
Lorenzo
L
Stella
Giampiero
G
Mei
Barbara
B
Orioni
Sarah
S
Ciccone
Giorgio
G
Federici
Mario
M
Lo Bello
Giorgio
G
Ricci
D005454:Fluorescence Polarization; D051549:Glutathione S-Transferase pi / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005982:Glutathione Transferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D011487:Protein Conformation; D055503:Protein Multimerization; D013050:Spectrometry, Fluorescence
C484555:GSTA1 protein, human; C496556:GSTP1 protein, human; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase; C412602:glutathione S-transferase Mu 2