Subunit dissociation

Polymer growth J(c) showed nonlinear monomer concentration dependence in the presence of ATP (Carrier et al., 1984), while in the presence of ADP, the plot of J(c) versus monomer concentration for actin was a straight line, as expected for reversible polymerization. The data imply that newly incorporated subunits dissociate from the filament at a slower rate than internal ADP-subunits in other words, (a) the effect of nucleotide hydrolysis is to decrease the stability of the polymer by increasing k and (b) nucleotide hydrolysis is uncoupled from polymerization and occurs in a step that follows incorporation of a ATP-subunit in the polymer. Newly incorporated, slowly dissociating, terminal ATP-subunits form a stable cap at the ends of F-actin filaments. 

Neuron-glial adhesion in nerve cell cultures is mediated by the pi subunit AMOG (adhesion molecule on glia) in the aipi isozyme of Na,K-ATPase [51]. Antibodies to the pi subunit dissociate cell-cell associations and also increase the rate of active... 

Binding of GTP promotes a disordering of the carboxyl- and amino-termini of the G-protein a subunit, with two parallel consequences the GTP-bound a subunit dissociates... 

Figure 6.3. Mechanism of action of heterotrimeric G-proteins. Upon receptor occupancy, the Ga-subunit binds GTP in exchange for GDP, and then moves in the membrane until it encounters its target enzyme, shown here as adenylate cyclase (alternatively, a phospholipase). The activated target enzyme then becomes functional. Inherent GTPase activity within the a-subunit then hydrolyses bound GTP to GDP, and the a-subunit dissociates from its target enzyme (which becomes inactive) and rebinds the / - and ysubunits. Upon continued receptor occupancy, further catalytic cycles of GTP exchange and target enzyme activation may occur. The scheme shown is for a stimulatory G-protein (Got,), but similar sequences of events occur with inhibitory G-proteins (Gcx,) except that the interaction of the a-subunit with adenylate cyclase will result in its inhibition. The sites of action of pertussis and cholera toxins are shown.

Cyclic AMP diffuses away from the membrane and engages its own target which is an inactive protein kinase, called cAMP dependent protein kinase or protein kinase A (PKA). The inactive PKA is a tetramer of two catalytic subunits and two regulatory subunits. Binding of cAMP to the regulatory subunits causes structural changes and the two regulatory subunits dissociate from the two catalytic sub-units. The now activated protein kinase A, that is the C subunit dimer, initiates a downstream cascade by... 

E. Casali, P. H. Petra, and J. B. A. Ross, Fluorescence investigation of the sex steroid binding protein of rabbit semm Steroid and subunit dissociation, Biochemistry 29, 9334-9343 (1990). 

Pressure may cause several changes in enzymes, as well as some changes which are not directly associated with the catalytic process. These changes may include conformational changes and subunit dissociation-association processes. Pressures above 4000 bar may induce conformational changes to such an extent that the enzyme in effect becomes irreversibly denatured. These are dealt with in the next section. In this section we will deal with lower pressures and reversible processes, namely, interactions between subunits in quaternary structures. For most multimeric enzymes, the maintenance of... 

In a pressure study involving a multimeric enzyme, it will in general not be possible to decide how much of the effect is due to direct influence of pressure on the catalytic process and how much of it is due to indirect influence through subunit dissociation and accompanying deactivation. Generally, a self-association reaction may be expressed in either of two equivalent forms ... 

From a study involving several multimeric and monomeric enzymes, Penniston (1971) concluded that subunit dissociation must be considered as the major determinant of the effect of pressure on enzymic systems. This view may not be generally accepted. 

Protein Subunit Dissociation-Association Volumes and Protein Denaturation Volumes ... 

Regulation of protein kinase A by cAMP takes place by the following mechanism. An increase in cAMP concentration, triggered by activation of adenylyl cyclase, leads to binding of cAMP at specific binding sites on the regulatory subunit. The R subunits dissociate from the tetramer, the catalytic subunits are released from inhibition by the regulatory subunits and can thus phosphorylate substrate proteins. 

Fig. 7.20. Regulation of glycogen-bound protein phosphatase I. Regulation of the activity of protein phosphatase I (PPI) takes place by phosphorylation of the G subunit. The G subunit is phos-phorylated at positions PI and P2, in the process of a signal chain mediated activation of protein kinase A. As a consequence of the phosphorylation, the catalytic subunit dissociates. The phosphatase activity of the free catalytic subunit is inhibited by association with a cytosohc protein phosphatase inhibitor (I), the binding of which is also controlled via a protein kinase A mediated phosphorylation. The phosphorylated G subunit can be dephosphorylated again by protein phosphatase 2A and may bind a catalytic PPI subunit once more.

FIGURE 5 Measuring [cAMP] with FRET. Gene fusion creates hybrid proteins that exhibit FRET when the PKA regulatory and catalytic subunits are associated (low [cAMP]). When [cAMP] rises, the subunits dissociate, and FRET ceases. The ratio of emission at 460 nm (dissociated) and 545 nm (complexed) thus offers a sensitive measure of [cAMP]. 

Denaturation. The stability of LADH and its denaturation has been studied under a variety of conditions including acid pH, and different concentrations of urea and guanidine hydrochloride. At pH 5, LADH loses its activity while still in the dimeric state, and at lower pH dissociation occurs, as can be seen in the drastic change in the fluorescence polarization spectrum.1410 The spectral data obtained are consistent with unfolding of the tertiary structure, some of which occurs before subunit dissociation. 

Frank M, Thumer L, Lohse MJ, Bunnemann M (2005)G protein activation without subunit dissociation depends on a Ga.-specific region. J Biol Chem 280(26) 24584-24590 Gao Z, Li BS, Day YJ, Linden J (2001) A3 adenosine receptor activation triggers phosphorylation of protein kinase B and protects rat basophilic leukemia 2H3 mast cells from apoptosis. Mol Pharmacol 59(1) 76—82... 

Frank, M., Thumer, L., Lohse, M. J., and Bunemann, M. (2005). G protein activation without subunit dissociation depends on a G alpha(i)-specific region./. Biol. Chem. 280, 24584-24590. 

Levitzki, A., and Klein, S. (2002). G-protein subunit dissociation is not an integral part of G-protein action. Chembiochem. 3, 815-818. 

Rebois, R. V., Warner, D. R., and Basi, N. S. (1997). Does subunit dissociation necessarily accompany the activation of all heterotrimeric G proteins . Cell. Signal. 9, 141-151. 

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