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Janson and Cleland 249 (8): 2562
J. Biol. Chem., Vol. 281, Issue 46, 37, November 17, 2006
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Classics
Determining Enzyme Mechanisms by Kinetic Analysis: the Work of W. Wallace Cleland
The Kinetic Mechanism of Glycerokinase
(Janson, C. A., and Cleland, W. W. (1974) J. Biol. Chem. 249, 2562–2566)
William Wallace Cleland was born in 1930 in Baltimore, Maryland. He received his A.B. from Oberlin College in 1950 and his M.S. and Ph.D. from the University of Wisconsin-Madison in 1953 and 1955, respectively. Cleland then did a postdoctoral fellowship at the University of Chicago, after which he returned to Madison to join the faculty of the University of Wisconsin as an Assistant Professor in 1959. He was promoted to Associate Professor in 1962, Professor in 1966, J. Johnson Professor of Biochemistry in 1978, and Steenbock Professor of Chemical Science in 1982. Cleland remains at the University of Wisconsin where he is still actively involved in research.
When Cleland first joined the faculty at the University of Wisconsin, his main focus was lipids. He spent several years studying the substrate specificity for acyl-CoA thioesters in the acylation of glycerophosphate to phosphatidic acids. He prepared acyl-CoA by chemical acylation of CoA, which at that time was only 75% reduced. Looking for a way to reduce the rest of the compound and keep it reduced, Cleland discovered that dithiothreitol was an excellent reagent for keeping thiols in the reduced state (1). His development of dithiothreitol as reducing agent for thiol groups resulted in the compound being named "Cleland's Reagent."
One of Cleland's duties as a young assistant professor was to teach a first year graduate level course in biochemistry. One of the topics on the syllabus was kinetics, and Cleland soon discovered that the field was in a very unsatisfactory state. He began to play with equations and carry out kinetic experiments on enzyme-catalyzed reactions. This led to his publication of three highly cited theoretical papers in Biochimica et Biophysica Acta (2–4) in which he proposed a nomenclature and theory for enzyme-catalyzed reactions with more than one substrate or product. These included reaction mechanism names such as "Ping-Pong Bi Bi."
Cleland used these methods to study various enzymes over the next decade or so. The Journal of Biological Chemistry (JBC) Classic reprinted here reports Cleland's analysis of the kinetic mechanism of glycerokinase (adenosine triphosphate:glycerol phosphotransferase), which catalyzes the following reaction.
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Using initial velocity and product inhibition techniques and dead-end inhibition by the intercoordination complex, chromium ATP, Cleland was able to determine that the reaction mechanism was Ordered Bi Bi with glycerol adding before the magnesium nucleotide and L-glycerol 3-phosphate being released last.
As he began to do more experimental kinetics, it became obvious to Cleland that statistical analysis of the data was needed, and he formulated equations and wrote several FORTRAN programs for applying the least squares method of curve fitting to enzyme kinetic data (5). Cleland also devoted a great deal of time to using isotope effects to study enzyme mechanisms, which is the subject of his 2003 JBC Reflections (6).
In recognition of his numerous contributions to science, Cleland has received many awards and honors. These include a 1986 Fulbright Senior Scholar Award, the 1990 Merck Award from the American Society for Biochemistry and Molecular Biology, the 1993 Alfred Bader
Award in Bioinorganic or Bioorganic Chemistry from the American Chemical Society, the 1995 Repligen Award from the Biological Division of the American Chemical Society, and the 1999 Stein and Moore Award from the Protein Society. Cleland has also served on the editorial boards of the Journal of Biological Chemistry and Biochemistry. He is a member of both the National Academy of Sciences and the American Academy of Arts and Sciences.
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REFERENCES
- Cleland, W. W. (1964) Dithiothreitol, a new protective reagent for SH groups. Biochemistry 3, 480–482[CrossRef][Medline] [Order article via Infotrieve]
- Cleland, W. W. (1963) The kinetics of enzyme-catalyzed reactions with two or more substrates or products. I. Nomenclature and rate equations. Biochim. Biophys. Acta 67, 104–137[Medline] [Order article via Infotrieve]
- Cleland, W. W. (1963) The kinetics of enzyme-catalyzed reactions with two or more substrates or products. II. Inhibition: nomenclature and theory. Biochim. Biophys. Acta 67, 173–187[Medline] [Order article via Infotrieve]
- Cleland, W. W. (1963) The kinetics of enzyme-catalyzed reactions with two or more substrates or products. III. Prediction of initial velocity and inhibition patterns by inspection. Biochim. Biophys. Acta 67, 188–196[Medline] [Order article via Infotrieve]
- Cleland, W. W. (1967) The statistical analysis of enzyme kinetic data. Adv. Enzymol. Relat. Areas Mol. Biol. 29, 1–32[Medline] [Order article via Infotrieve]
- Cleland, W. W. (2003) The use of isotope effects to determine enzyme mechanisms. J. Biol. Chem. 278, 51975–51984
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