Enzyme dissociation constant

Lineweaver, H. and Burk, D. (1934) The determination of enzyme dissociation constants. Journal of the American Chemical Society, 56, 658-666. 

Lineweaver, H. and Burk, D. (1934) Determination of enzyme dissociation constants./. Am. Chem. Soc., 56, 658. Eadie, G.S. (1942) The inhibition of cholinesterase by physostigmine and prostigmine. /. Biol. Chem., 146, 85. Hofstee, B.H. (1959) Non-inverted versus inverted plots in enzyme kinetics. Nature, 184, 1296. 

No discussion of this subject would be complete without emphasizing the point that the relative rates of hydrolysis of two substrates, which may be quite different from their relative affinities for an enzyme, may be at least as important as regards the physiological action of the enzyme. Dissociation constants measure affinity only (affinity = 1/dissociation constant). Thus, despite its 10-fold smaller affinity, phenyl /3-glucuronide is hydrolyzed nearly as rapidly as phenolphthalein /3-glucuronide by the purified enzyme from female-rat preputial gland at saturation with both substrates, in terms of moles of aglycon liberated.40 In the presence of a combined inhibitor (see Section IX, 3), there is a fall in the rate of hydrolysis, despite the increased affinity of the enzyme for the substrate. 

The active site structure of trypsin-like enzymes is considered to be very similar to that of bovine trypsin, yet little is known about them. Refinement of these structures is important also for the purpose of designing physiologically active substances. With a view to comparing the spatial requirements of active sites of these enzymes, dissociation constants of the acyl enzyme-ligand complex, K-, which were defined before, were successfully analyzed By taking advantage of inverse substrates which have an unlimited choice of the acyl component, development of stable acyl enzymes could be possible. These transient inhibitors for trypsin-like enzymes could be candidates for drugs. In this respect, the determination of the deacylation rate constants for the plasmin- and thrombin-catalyzed hydrolyses of various esters were undertaken 77). 

A dissociation constant is a constant whose numerical value depends on the equilibrium between the dissociated and undissociated forms of a molecule. Higher the dissociation constant, greater the dissociation. Examples of dissociation constants are substrate-enzyme dissociation constant and the acid dissociation constant pfC. This latter is defined as... 

This implies that every collision of a reactant molecule (substrate) with the enzyme leads to product formation. Because tor many enzymes (the Michaelis or apparent substrate-enzyme dissociation constant) values are in the micro- or submiaomolar range (10 M), one can compute the (the catalytic or... 

Km, asmaybe seen from equations (1) and (6), is the dissociation constant of the enzyme-substrate compound (JES) and has a characteristic value for each enzyme. It is known as the Michaelis constant, and the reciprocal /Km = Km, is termed the association constant. Equation (12) has been used extensively for the calculation of enzymic dissociation constants, but the method has been much improved by Lineweaver and Burk (S), who employ the reciprocal of equation (11) for the calculation ... 

It is obvious that 1/k instead of (l/k )V may be employed since Km is determined under such conditions that both quantities are zero. This equation is probably the most convenient form to use for the determination of enzyme dissociation constants. This form was first suggested by Veibel U) > but the development of the equation as given here is original. 

Cytochrome c derived from tuna fish reacts slower with the immobilized oxidase compared to cytochrome c derived from horse at the same concentration. This is due to a smaller protein/enzyme dissociation constant for tuna protein compared to horse protein. These data show that electron transfer from tuna cytochrome c to the enzyme is rate limiting even at concentrations of 10 xM and above. 

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