Enzymes volume change

Figure 12.3 Rigid and flexible molecular interaction field maps with the hydrogen probe in the active-site cavity for CYP2C9 and UCT2B7 enzymes. It is worth noting that, with flexible side chains, the overall cavity volume changes considerably. This demonstrates the important role played by MIF-flexibility calculations in enzyme-substrate recognition.

Under specific conditions one may be able to distinguish the last two phases and separate their volume contributions, whereas under other conditions one may not. By means of special techniques and with very simple enzymes it may even be possible to separate volume changes within one single phase. However, with ordinary techniques and especially without a thorough theoretical treatment, the measured volume changes will be mixtures of all the above-mentioned contributions. 

Related to this is the volume change associated with dipole development in transition states. This has been investigated theoretically for a model substance of molecules with a size similar to that of water (Morild and Larsen, 1978). The calculations show that the volume changes are very pressure-dependent in this case. A change in dipole moment from 0 to 1 x 10-3 C-m gives a volume decrease of about 30 cm3 mol-1 at 350 bar and about 20 cm3 mol-1 at 750 bar. However, this may not be typical for molecules as large as enzyme-substrate complexes. 

The volume phase transition temperatures Tc for collapsing and Ts for swelling changed depending on the activity of the entrapped enzyme. When the enzyme was active in the presence of 47.3 mM of ethyl butyrate, Tc = 29.8 °C and Ts = 29.1 °C, while Tc = 28.4 °C and Tg = 27.6 °C with inactivated enzyme. The change in the phase transition temperature was presumably caused by the change in the substrate and product composition induced by enzymatic reaction within the gel phase. 

AV means the volume difference of the activated complex and of the educts which are the substrates and the free enzyme. A difference in the volumes of the activated complex and of the educts leads to an increase or decrease of the reaction rate depending upon a positive or negative volume change. According to Chen the enantioselectivity is defined as... 

Considering that the enantioselectivity (E) is dependent on high hydrostatic pressure the KM and v value should also be dependent on pressure. In view of the enzyme catalysed reactions volume changes due to conformational changes of the enzymes should expect larger sensitivity of the reaction rates compared to the uncatalysed ones. 

All of the preceding remarks apply equally well to the studies which have been made of the effects of high pressures on the rate constants of enzyme reactions and also protein reactions. The values of AFf and AF which can be obtained from the pressure coefficients of the rate constants cannot be naively interpreted in terms of simple volume changes of the enzyme or protein without a careful assessment of the other parameters of the system and their changes with pressure. ... 

Fig. 3 Sample ENPP1 enzyme dependence. Change in 0D635 values are plotted against ENPP1 concentration in 6 il assay volume, and the data fitted linearly. Slope is 0.68 0.05, standard deviation of the background control is 0.0071, and the limit of detection (LOD) is 0.031 nM...

High hydrostatic pressure alters any process that proceeds with a volume change. This affects enzyme catalysis because both substrate binding and associated protein conformational changes proceed with changes in the hydration state of amino acid residues that alter the total volume of the water-protein-ligand system (Low and Somero, 1975). Therefore, animals adapted to the deep sea require changes to the amino acid composition of enzymes that create pressure-insensitive kinetic parameters and pressure-resistant structures. 

The picture arrived at after exhaustive research with luminous bacteria, and eventually discussed at length with reference to many other types of biological processes, involved molecular volume changes of activation in the enzyme reaction and volume changes of reaction in the reversible denaturation of the protein moiety. Continued research concerning the... 

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