Lineweaver-Burk analysis

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. 

In a Lineweaver-Burke analysis Mk pp is plotted as a function of l/[catalyst]. The intercept of this plot on the y-axis provides the value of Vk-yA, whereas the intercept on the x-axis yields the value for 1/K,z. 

Schultz s group employed an a-hydroxyphosphonate hapten [99] and subsequently isolated 20 cell lines of which 5 catalysed the hydrolysis of the model substrate p-nitrophenyl phosphate [100] above background (Fig. 34) (Scanlan et al., 1991). Antibody 38E1 was characterized in more detail and kinetic parameters were afforded by Lineweaver-Burke analysis. This antibody exhibited 11 turnovers per binding site with no change in Vmax, and thus acted as a true catalyst. Moreover, examination of substrate specificity showed that catalysis was entirely selective for p-substituted species (Appendix entry 6.6). 

Kinetics of Free and Mao-osoib-Bound eaiymc. The results of the Lineweaver-Burk analysis of the initial rates for free and immobilized enzyme appear in Table I. The increase in the substrate affinity parameter due to some internal mass transfer limitations in the IME as no substrate-matrix interactions were present. The increase... 

Lineweaver-Burk analysis using the substrate saturation curves afforded the corresponding Michaelis-Menten kinetic parameters of the reaction V max=l-79 xIO- Ms , KM=21.5mM, kcat = 8.06x 10 s for 69, and Knax = 9.22x 10... 

The representative example, MC 52201 80 was kinetically well behaved, proving to be a classical competitive, time-independent PTP1B inhibitor after a Lineweaver-Burk analysis. Further studies on this class are reported as on-going. 

Reversible inhibition can be competitive or non-competitive. Competitive inhibitors bind to the active site and compete with the substrate for binding to the enzyme. However this means that increasing the S concentration will progressively outcompete the inhibitor. Accordingly a Lineweaver—Burk analysis of enzyme kinetic data obtained in the presence or absence of a competitive inhibitor will yield the same Fmax (at infinite S concentration) but the Am in the presence of the inhibitor (A, ) will be higher (poorer binding) than the Am measured in the absence of competitive inhibitor. Knowing the inhibitor concentration [I] one can calculate the A) from the relation ... 

Figure 9-3. Error propagation in the direct analysis and Lineweaver-Burk analysis of Michaelis-Menten kinetics.

Since the nonlinear least-squares method requires initial guesses to start the procedure, three different initial trials were performed (1) (0,0), (2) (1,1), and (3) the values obtained from the Lineweaver-Burk plot in Example 4.2.4. All three initial trials give the same result (and thus the same relative error). Note the large differences in the values obtained from the nonlinear analysis versus those from the linear regression. If the solutions are plotted along with the experimental data as shown below, it is clear that the Lineweaver-Burk analysis does not provide a good fit to the data. 

Lineweaver-Burke analysis of the steady-state kinetic data for A5 revealed a k =0.l min equivalent to a rate enhancement of 290. The background reac-... 

The Lineweaver-Burk analysis yields slope Kl vm and intercept l/vm. Use the next two rows to convert this information into the parameters of interest, Kand vm. Here, K slope / intercept, and vm= intercept. 

A detailed study of the mechanism of APTl inhibition by the palmostatins was performed [7]. Lineweaver-Burke analysis revealed that with varying inhibitor concentration is increased and remains constant, which indicates... 

A cocrystal structure of the protein kinase Pim-1 and 1 showed that the complex 1 binds to the ATP binding site in a homologous manner to staiu-osporine (Figure 1.3) [21]. Previously, a Lineweaver-Burke analysis, which is a kinetic... 

Low concentrations of carba-NAD have been demonstrated to inhibit the NAD glycohydrolase isolated from Bungarus fasciatus venom (11). The mechanism of inhibition was determined by Lineweaver-Burk analysis of the effect of fixed inhibitor concentration on the initial rate of the reaction (Fig. 4). The inhibition of the NAD glycohydrolase by mixtures of [1] and [7] was determined to be of the competitive type. Replots of the KmW versus inhibitor concentration (not shown) were linear, and a Ki of 74 pM determined. It is our expectation that the Ki for pme carba-NAD will be lower than this value. As it stands, the Ki for carba-NAD is close to the Km of NAD, which we determine to be 33 pM. 

Nitric oxide synthase (EC 1.14.13.39) incubated with both doxorubicin and menadione augmented the rate of NADPH oxidation (Garner et al. 1999). Kinetic parameters by lineweaver-Burk analysis showed that NOS III had the highest affinity for both doxorubicin and menadione with K s of 40.6 and 22.7 pM, respectively. NOS II had the lowest affinity for both compounds with K s of 210.5 and 44.1 pM, respectively. 

The fact that catechol oxidation occurs via a Michaelis-Menten mechanism is confirmed by obtaining the RDE response for a range of catechol concentrations and subsequently subjecting the data to a Koutecky-Levich analysis [see (Fig. 2.36(a)]. The Koutecky-Levich intercept 7/cz, is then obtained and a Lineweaver-Burk analysis applied, which predicts that //fi, varies in an inverse manner with bulk substrate concentration according to... 

FIGURE 2.36. (a) Koutecky-Levich plots for catechol oxidation in 0.2 M H2SO4 at the metal oxide/ionomer composite electrode as a function of catechol concentration, (b) Lineweaver-Burk analysis of data in Fig. 2.36(a). 

Inhibition Constants In inhibition experiments, dataware collected for 4 to 5 different substrate concentrations at each of 4 to 6 different inhibitor concentrations (ranging from zero added inhibitor, to sufficient inhibitor to reduce the observed activity by approximately 80%). Lineweaver-Burk plots of the resulting data were analyzed by unweighted linear least squares analysis, yielding a slope and ordinate intercept at each inhibitor concentration. (Due to its prevalence in the literature, Lineweaver-Burk analysis was the analytical method of choice. Because of the magnitude of error in our data was independent of velocity, and... 

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