Liver hydroxymethylbilane synthetase (EC : an environmental and toxilogical kinetic study. The Effect of experimental design and circumstance on three common linearization methods used in determining the Michaelis-Menten kinetic constants


Farmer, Douglas John




Hydroxymethylbilane Synthetase (EC was partially purified from rat liver and spleen, human erythrocytes, common gull liver and ring dove liver by stepwise centrifugation and heat denaturation of interfering proteins. The kinetics were found to be unimolecular and consistent for the rat tissues and human erythrocytes and gave K. values of between 6 and 12 μM for the rat tissues and 5 and 8 μM for the human RBC. The dove and gull tissues proved to be less consistent and gave K. values of between 5 and 6 μM for the dove tissues and 3 and 10 μM for the gull tissues. Enzyme for doves proved to be sensistive to various organic toxins and showed definite response during dosings with lead acetate and trichlorobenzene as well as benzene, carvone, benzanthracene, phenobarbital and lindane. The rat liver enzyme proved sensitive to a large spectrum of meavy metal ions and indicated the possible existence and cytsteine, histidine and tryptophan at the active site.

Computer simulations were done to determine the effects of experimental error and design on the three common kinetic linearization methods. It was found that the plot of Lineweaver and Burke was the most accurate in determining these constants while that of Haines and Woolf proved to be the best at indicating poor data. In addition running kinetic experiments at two substrate concentrations as opposed to ten was found to give better estimates as did making more velocity determinations at the lowest velocity.


Hydroxymethylbilane Synthetase




Carleton University

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