Course Syllabus

Biological Chemistry/Chemistry 673: Mechanism & Kinetics of Enzymes  Broadcast from 1632 Chemistry, MWF 11:00 - 11:50

Kinetics for Enzymologists? Hell yeah - Laissez les bons temps rouler!!!

This course delivers an in-depth introduction to the chemical kinetics that describe enzyme action with the goal of developing the skills needed to elucidate enzyme mechanisms. Starting with simple unimolecular reactions, paradigms of increasing complexity are considered along with transient kinetic experiments used to analyze them. This march through key mechanisms reaches a crescendo when we consider catalytic cycles from the point of view of transient kinetics. We then switch viewpoints and analyze steady-state kinetic mechanisms, leaning heavily on Cleland's nomenclature and rules for the analysis of steady-state velocity patterns. We close by considering the complementary information that transient kinetics and steady-state kinetics provide to the mechanistic enzymologist.

While the riveting lectures develop the theory behind kinetic analyses, both mathematically and qualitatively, the rubber meets the road in the frequent problem sets. You'll put theory into practice with virtual experiments, and you'll practice your theory with derivations and simulations. An emphasis is placed on learning to design and interpret experiments, analyzing new kinetic situations, and using research-grade software.

It's intense, and it's great fun. Biol. Chem. 673 - the toughest course you'll ever love.

 

Useful texts:

No textbook adequately covers the topics in this course, and, therefore, none is required. A recent book comes closest: Clive Bagshaw, Biomolecular Kinetics: A Step-By-Step Guide, 2017, CRC Press, Taylor & Francis Group, Boca Raton. The newest book on the scene organizes things differently from 673 but is a solid treatment of the subject, relying heavily on simulations using KinTek Explorer: Kenneth A. Johnson, Kinetic Analysis for the New Enzymology, 2020, KinTek Corp. Also, these two books may provide useful perspectives: Alan Fersht, Structure and Mechanism in Protein Science, 4th ed., 2017, World Scientific Press (essentially the same as the 1999 3rd ed.); and, Paul F. Cook & W. W. Cleland, Enzyme Kinetics and Mechanism, 2007, Garland Science, London and New York

 

Software: Many of the exercises in this course will require:

• Kaleidagraph is strongly recommended for curve-fitting. (No, you can't use Excel instead!! And yes, K'graph is better than Prism.) A free demo version that lasts 75 days is almost fully functional. It may be obtained at: http://www.synergy.com  The licensed version is found on the computers of many research groups on campus, or, buy it yourself – it's cheaper than many textbooks. Important for Mac-users: Apparently Kaleidagraph does not work yet for Catalina (OS X 10.15). If you haven't upgraded to 10.15 yet, please delay! If you have, please contact me.

 

• For simulating mechanisms, we will use Berkeley Madonna, a research-grade program for solving differential equations (and doing other things): https://berkeley-madonna.myshopify.com   We have purchased year-long licenses for 673 (codes to be distributed by email). To download a version of Madonna that accepts our license-codes, use this link for Mac and this link for Windows. Important for Mac-users: Apparently Madonna does not work yet for Catalina (OS X 10.15). If you haven't upgraded to 10.15 yet, please delay! If you have, please contact me. 

 

• We will also have temporary licenses partway through the course for KinTek Explorer, a global fitting program with an unusually friendly interface: https://www.kintekexplorer.com  . Important for Mac-users: Apparently KTE does not work yet for Catalina (OS X 10.15). If you haven't upgraded to 10.15 yet, please delay! If you have, please contact me. 

 

• Virtual experiments will be performed using webZyme. Your user name will be your unique name, and your default password will be "webzyme".

 

Grading:

Grades will be assigned on the basis of problem sets (high-value) and surprise quizzes (low-value).

 

Learning Objectives/Outcomes:

By the end of this course, you will have an advanced understanding of how to elucidate enzyme mechanisms using transient and steady-state kinetics. You will be able to design incisive experiments, use research-grade software to analyze data, build kinetic models, and test these hypotheses with new experiments. You will be able to critically analyze your own experiments and work in the literature.

 

Tentative Schedule:

W	8 Jan	Organizational Issues; Basic Concepts; Rates, Rate Constants, & Mass Action
F	10 Jan	Numerical & Analytical Solutions; First-Order Irreversible
M	13 Jan	First-Order Irreversible
W	15 Jan	First-Order Reversible; Kinetics & Equilibrium Constants
F	17 Jan	Bimolecular Irreversible; Pseudo-First-Order Approximation
M	20 Jan	No Class – MLK day
W	22 Jan	Diffusion Control
F	24 Jan	Bimolecular Reversible
M	27 Jan	Bimolecular Reversible; Ligand-Binding
W	29 Jan	Titrations
F	31 Jan	Competition
M	3 Feb	Two-Step Sequential; Rate-Determining Steps
W	5 Feb	Spectral Ambiguity; N-Step Reactions
F	7 Feb	Spectral Ambiguity; N-Step Reactions; Spectra of Intermediates
M	10 Feb	Spectra of Intermediates
W	12 Feb	Reality-Check 1 – A Nice Example
F	14 Feb	Reality-Check 1 – A Nice Example
M	17 Feb	Reality-Check 1 – A Nice Example
W	19 Feb	Two-Step Reversible – Rapid-Equilibrium Approximation; Ligand-Binding; pH Effects
F	21 Feb	Two-Step Reversible – Rapid-Equilibrium Approximation; Ligand-Binding; pH Effects
M	24 Feb	Two-Step Reversible – Steady-State Approximation; Application to Ligand-Binding
W	26 Feb	Two-Step Reversible – More Ligand-Binding
F	28 Feb	Two-Step Reversible – More Ligand-Binding; Competition
M	2 Mar	No Class – Spring Break
W	4 Mar	No Class – Spring Break
F	6 Mar	No Class – Spring Break
M	9 Mar	Two-Step Reversible – Exact Solution
W	11 Mar	Two-Step Retrospective; N-Step Reversible
F	13 Mar	Reality Check 2 – Another Nice Example
M	16 Mar	Reality Check 2 – Another Nice Example
W	18 Mar	Turnover
F	20 Mar	Bursts
M	23 Mar	Transient Kinetic Retrospective
W	25 Mar	Types of Experiments
F	27 Mar	Steady-State Kinetics – Cleland's Nomenclature & Diagrams
M	30 Mar	Steady-State Kinetics – Net Rate Constants
W	1 Apr	Steady-State Kinetics – King-Altman
F	3 Apr	Steady-State Kinetics – Kinetic Mechanism from Patterns
M	6 Apr	Steady-State Kinetics – Inhibition; Kinetic Mechanism from Patterns
W	8 Apr	Steady-State Kinetics – Inhibition; Kinetic Mechanism from Patterns
F	10 Apr	Steady-State Kinetics – Consistency with Transient Kinetics
M	13 Apr	Steady-State Kinetics – Consistency with Transient Kinetics
W	15 Apr	Building Complex Kinetic Models
F	17 Apr	Building Complex Kinetic Models; Isotope Effects
M	20 Apr	Global Analysis; Retrospective

 

 

Prerequisites: Senior-level biochemistry and undergraduate calculus are highly recommended.

 

Attendance: Attendance is optional, but miss class at your own risk! There is no other source for this material organized in this way, and each topic builds on previous topics.

 

Group-work: Many of the homework problems are challenging. Sometimes it helps to work in groups to brainstorm. This is encouraged, but beware! Sometimes groups become enthralled with poor ideas; you must think for yourself. Also note that each student has customized webZyme exercises, so you'll need to do you own work even if you establish your thinking by working in a group.

 

Deadlines: Homework is due in class one week after the assignment is made, unless stated otherwise. Extensions might  be granted upon request. Surprise quizzes are given in the first five minutes of class. There are no make-up quizzes.

 

Tips for Success:   Many students think 673 is hard. This course does not need to be as hard as some students make it. In order to do well with minimal pain:  

1. Ask questions immediately if anything is ever unclear in class. New material builds on the previous material; you must understand it as we go. You should not wait to figure it out later; that causes unnecessary suffering.  

2. Review your notes regularly. Because there are no exams, the incentive to review older material is less apparent. This is a mistake. No old material becomes irrelevant, and at the end of the course you'll need to grab insight developed throughout the whole course.  

3. Don't wait until the last minute to start your homework. Homework problems generally require some contemplation and/or extensive (virtual) experimental analyses, and then thoughtful interpretation. Maybe sometimes even creativity. These take a little time.

4. Before you plunge ahead in attacking a homework problem, pause briefly and ask yourself why you're about to do what you're about to do. If your answer amounts to "I dunno, I think that's just what you're supposed to do," then stop. Do things because they make scientific sense.

 5. Ask me if you have questions about your problem-solving strategies. The point of the homework problems is for you to learn, not to trick you out of points.  

6. If you're having trouble with the software, send me an email immediately, describing the problem in detail. It's often helpful to know what your operating system and web-browser are.