About the author: Kenneth A. Johnson is The Roger Williams Centennial Professor of Biochemistry at The University of Texas at Austin, and President and founder of KinTek Corporation, Austin, Texas, USA. He previously served as the Paul Berg Professor of Biochemistry at The Pennsylvania State University. His current research seeks to define the kinetic, thermodynamic, and structural basis for the extraordinary specificity of DNA polymerases, and to apply that knowledge to improve treatments for viral infections.


This book was written for anyone who wants to understand the principles and practical use of kinetic and equilibrium methods to define reaction pathways, in particular, graduate students and postdoctoral fellows in training as well as academic and industrial research scientists. 


Traditionally, enzymology has been centered on the use of steady-state kinetic methods, which represent a good beginning but are limited in their ability to define the mechanisms of catalysis. In the New Enzymology, we shift our focus beyond the steady state to experiments that afford direct observation of reactions occurring at the enzyme active site. More importantly, this book presents a marked departure from the way in which kinetics has been taught in the past. Rather than deriving equations for different models and then learning to choose the appropriate equation for each experiment, we focus on the use of computer simulation with fast algorithms to model experiments as they are performed and without simplifying approximations. We work to develop a fundamental understanding of reaction kinetics and equilibria through hands-on experience via simulation of numerous enzyme systems and experimental approaches. %Reaction mechanisms are derived by globally fitting data collected using different approaches to examine the reactions.


This book is built around the use of KinTek Explorer software to teach kinetic principles and methods of data fitting. KinTek Explorer is available for Windows and Mac OS at https://www.kintekexplorer.com. For all of the exercises given in this book, the software can be run in student mode without requiring a paid license. This software is structured to mimic the thought process involved in designing an experiment, so learning to use the software translates into learning how to design and interpret effective experiments.  Another key feature of KinTek Explorer is dynamic simulation, a process where you can scroll the value for a given concentration, rate constant, or output scaling factor while simultaneously observing changes in the output signal. This process provides a visual link between experiments and models to learn kinetics and allows you to explore the range over which parameters can be varied while fitting data. In each chapter, simulation exercises are provided to illustrate principles underlying observable kinetics. 


Although equations and their derivations are presented, and they are useful in illuminating patterns in the data, fitting data using equations has serious limitations that are overcome by the use of computer simulation. More importantly, rather than fitting data in a piecemeal fashion and then trying to put the pieces together, we now fit multiple experiments simultaneously using a single unifying model to account for all experimental observations. This represents another marked departure from traditional methods of data fitting. We illustrate global data fitting with numerous examples that demonstrate that the whole is greater than the sum of the parts. Although most examples rely on enzyme kinetics, the analysis presented here applies equally well to topics ranging from chemical kinetics to pharmacokinetics/dynamics and metabolic pathways. 


The chapters are organized to logically build knowledge and conceptual understanding. However, much of what is presented is interconnected, so it is difficult to present the material linearly. For example, we start with equilibrium binding measurements in Chapter 3, but to understand equilibria we need to include reference to the kinetics of ligand binding that are not presented until Chapter 8. An argument could be made to postpone discussion of steady-state kinetics (Chapter 5) until after covering the kinetics of single reaction steps during turnover (Chapters 8 to 10), but most students will have already been introduced to steady-state kinetics, so we retain an order that follows the historical progression in enzymology. In addition, each chapter contributes to building an understanding of data-fitting through examples, but we wait until Chapter 15 to provide an in-depth discussion of the statistical analysis. Our purpose is to retain focus on enzymology rather than statistics, but at the same time we provide a strong foundation as needed. To facilitate a balanced presentation, we include extensive cross-referencing between chapters. When reading a given chapter, you can choose to ignore references to a different section to maintain continuity in your reading, or follow the lead if your curiosity or desire for clarification needs to be satisfied. 


Several chapters are dedicated to discussing examples of kinetic analysis to solve mechanistic questions. These chapters illustrate the application of principles developed in the preceding chapters in an attempt to bring the concepts to life. Although you may not care specifically about the mechanisms of HIV reverse transcriptase, EPSP synthase, tryptophan synthase, or dihydrofolate reductase, discussion of the design and interpretation of the experiments are an integral part of the instruction. The examples are intended to stimulate you to think about your own research and how similar experiments could be applied to your system.  


This book and KinTek Explorer are the culmination of 40 years of research by the author on scores of enzymes, and they are offered with the sincere desire to pass on the insights learned from each enzyme and with the hope that you will delight in discovering the beauty and power of kinetic analysis.