Effect of polyethylene glycol on the activity, intrinsic fluorescence, and oligomeric structure of castor seed cytosolic fructose‐1, 6‐bisphosphatase

Abstract

The effect of polyethylene glycol (PEG) on the activity, intrinsic fluorescence, and oligomeric structure of homogeneous cytosolic fructose-l,6-bisphosphatase (FBPasec) from endosperm of germinating castor oil seeds has been examined. Increasing the PEG concentration in the FBPasec reaction mixture elicited a progressive 3-fold decrease in the enzyme's K~ for fructose-l,6-P 2. The presence of PEG also: (i) increased the extent of FBPase c inhibition by high levels of fructose-l,6-Pz, (ii) enhanced the intensity of the enzyme's fluorescence emission spectra, and (iii) prevents dissociation of the active tetrameric native enzyme into inactive lower Mr forms during gel filtration HPLC. It is concluded that the activity and structure of plant FBPase c is modified by extreme dilution, probably as a result of partial deaggregation of the native tetrameric enzyme.

Kinetic and structural analysis of the ultrasensitive behaviour of cyanobacterial ADP-glucose pyrophosphorylase

“…Indeed, in the absence of PEG, the enzyme behaved as a 200 kDa tetramer, as could be judged by gel-filtration chromatography on a Superose 12 column (Figure 6A), and retained the same quaternary structure, despite the presence of its substrates or allosteric effectors (results not shown). In agreement with previous reports [14], we observed that the Superose 12 column was unable to separate the different standard proteins when chromatographed in the presence of PEG. To solve this inconvenience, we used a Sephacryl S-300 column for running in the presence of the inert polymer (Figure 6B).…”

“…In crowded media, protein aggregation is promoted through its exclusion from the solvent and the consequent local increase in protein concentration [7,13]. The kinetic and structural properties of oligomeric proteins may be affected by both homologous and heterologous protein-protein interactions [8,14]. Previous experimental evidence exists which shows that the presence of PEG increases the activity of a variety of enzymes [12,15].…”

Effects of Macromolecular Crowding on Glycoprotein Processing Enzymes

“…Under these circumstances, compact conformation of macromolecules is favored, and their association would be enhanced . As a result, elevation of enzyme activities has been observed in certain cases. − Such phenomena are referred to as volume exclusion effect (positive macromolecular crowding effects) . However, a crowded environment decelerates the diffusion of molecules, and the rate of diffusion-controlled reactions may decrease (negative macromolecular crowding effect) .…”