Lineweaver-Burk Method

Lineweaver—Burk Method.8 The Michaelis-Menten equation may be algebraically rearranged to Eq. 2.22, yielding a third linear plot, called the Line-weaver-Burk or double-reciprocal plot  

The reciprocal rate 1/v is plotted against the reciprocal substrate concentration 1/ [S], yielding a straight line with a positive slope of Km/Vmax and a y intercept of 1/Vmax- The parameter Km is then calculated from the linear regression values of the slope divided by the y intercept (Fig. 2.9). While the Lineweaver-Burk plot is 

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In each of these cases, the double-reciprocal plots according to the Lineweaver-Burk method are linear. The appearance of these plots, and the parameters obtained from them, are developed in Problem 4-15. 

The kinetic parameters (Km and l/max) were determined through the conventional Lineweaver-Burk method, by varying the sucrose concentration between 10.0 and 292 mM. 

The initial rate of uptake also increases as the external concentration is raised (Figure 2). At high concentrations further increments in the extracellular concentration produce much smaller increases in the accumulation rate, indicating the involvement of a saturable component in the uptake process. At low concentrations the data conform to the classical equation of Michaelis-Menten, giving a straight line when plotted according to the Lineweaver-Burk method. The rising slope of... 

Henderson-Hasselbalch equation lahn-Teller effect Lee-Yang-Parr method Lineweaver-Burk method Mark-Houwink plot Meerwein-Ponndorf theory Michaelis-Menten kinetics Stem-Volmer plot van t Hoff-Le Bel theory Wolff-Kishner theory Young-Laplace equation Ziegler-Natta-type catalyst... 

The Lineweaver-Burk Method This method uses the rearranged equation... 

Pig. (first) The Lineweaver-Burk method of plotting enzyme kinetic data (second) Eadie-Hofstee method of plotting enzyme kinetic data. 

Thus, the Lineweaver-Burk method describes the behavior of (rate) but not (rate). 

This example illustrates how the nonlinear least-squares method can be used and how initial guesses must be explored in order to provide some confidence in the solution obtained. It also demonstrates the problems associated with the Lineweaver-Burk method. 

A plot of v (as y) against v/[S] (as x) will yield, after linear regression, a y intercept of Emax and a slope of A nl (Fig. 2.7). This plot is the preferred linear regression method for determining Km and V/ln ix, since precision and accuracy are somewhat better than those obtained using the Hanes plot, and much better than those found using the Lineweaver-Burk method. 

Lineweaver-Burk method yields significantly poorer precision than any of the other three methods. The Comish-Bowden-Eisenthal method clearly yields the most precise values for Km, in the presence of either absolute or relative error. 

Two experiments were performed with the enzyme ribonucle-ase. In experiment 1 the effect of increasing substrate concentration on reaction velocity was measured. In experiment 2 the reaction mixtures were identical to those in experiment 1 except for the addition of 0.1 mg of an unknown compound to each tube. Plot the data according to the Lineweaver-Burk method. Determine the effect of the unknown compound on the enzyme s activity. (Substrate concentration is measured in millimoles per liter. Velocity is measured in the change in optical density per hour.)... 

Fig. 3.5 Spreadsheet for the analysis of the data on the kinetics of nicotinamide mononucleotide adenyl transferase by Atkinson etal, Biochem.J. 80 (1961) 318. The results should be read as, e.g., on line 30 K = 0.4406 0.0906 for the unweighted Lineweaver-Burk method, K = 0.5821 0.0522 for the unweighted Hanes plot, or, on line31, vm = 0.5853 0.1039 for Lineweaver-Burk, vm= 0.6848 0.0383 forHanes.

Henderson—Hasselbalch equation Jahn—Teller effect Lineweaver—Burk method Mark-Houwink plot Meerwein—Ponndorf theory Michaelis—Menten kinetics Stern—Volmer plot van t Hoff—Le Bel theory Wolff—Kishner theory Young—Laplace equation Ziegler—Natta-type catalyst... 

Sucrose (common table sugar) is hydrolyzed to glucose and fructose (Section 16.3) in a classic experiment in kinetics. The reaction is catalyzed by the enzyme invertase. Using the following data, determine, by the Lineweaver-Burk method, whether the inhibition of this reaction by 2 M urea is competitive or noncompetitive. 

To overcome the limitation of the Lineweaver-Burk method, another graphic method for and determination has been proposed, where linearization is achieved by the mnltiplic ipn of both sides of Equation 4.23 by [5], as shown... 

The Lineweaver-Burk method is primarily used nowadays for initial examination of kinetic data, and then the data are fitted to an appropriate rate equation using computer programs. 

The equilibrium constants obtained using the metal-ion induced shift in the UV-vis absorption spectrum are in excellent agreement with the results of the Lineweaver-Burke analysis of the rate constants at different catalyst concentrations. For the copper(II)ion catalysed reaction of 2.4a with 2.5 the latter method gives a value for of 432 versus 425 using the spectroscopic method. 

Lineweaver-Burk plot Method of analyzing kinetic data (growth rates of enzyme catalyzed reactions) in linear form using a double reciprocal plot of rate versus substrate concentration. 

Methods of plotting data obtained from enzyme catalyzed reactions, (a) Lineweaver-Burk plot, (b) Eadie or Hofstee plot, (c) Hanes plot. 

Lineweaver-Burk plot (Figure 8.11). In such cases the chosen substrate concentration must give the highest reaction velocity possible. It is important, when describing any enzyme assay, to report the percentage maximum velocity which the method will give. 

INDIVIDUAL REACTION MECHANISMS LINEWEAVER-BURKE PLOT POINT-OF-CONVERGENCE METHOD SCATCHARD PLOT... 

Since k0 is known, Equation (29) allows km to be evaluated. Likewise, the equilibrium constant for the binding of the substrate to the micelle can be evaluated from Equation (31) if n is known from a separate experiment. This method of analysis of catalyzed reactions is called a Lineweaver-Burke plot after the corresponding technique in biochemistry. Example 8.5 illustrates the use of these relationships. 

The Lineweaver-Burk plot of the experimental data is shown in Figure 3.7. The straight line was obtained by the method of least squares. From the figure, the values of and were determined as 2.4gmol m and 7.7 x 10 ... 

It is very useful to transform the Michaelis-Menten equation into a linear form for analyzing data graphically and detecting deviations from the ideal behavior. One of the best known methods is the double-reciprocal or Lineweaver-Burk plot. Inverting both sides of equation 3.1 and substituting equation 3.2 gives the Lineweaver-Burk plot 4... 

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