Proteins carry out most functions in the cell. These molecules are not rigid, but are mobile on a wide range of timescales. The course aims to convey the concept of protein dynamics in relation to biological function. The focus is on enzyme catalysis and two techniques will be discussed in detail: nuclear magnetic resonance (NMR) spectroscopy to study protein mobility and kinetic measurements to determine enzyme activity. NMR is a versatile analytical technique with many applications. We will discuss the fundamentals of NMR (chemical shift, dipolar coupling, J-coupling, relaxation), as well as two-dimensional NMR and relaxation experiments to characterize molecular mobility on nanosecond and millisecond timescale. It provides a basis in NMR spectroscopy that is of use for any chemist. The study of kinetics will comprise steady state and pre-steady state theory and methods to measure enzyme characteristics such as kcat and KM, as well as inhibitor kinetics.
Lectures, exercises, demonstrations and simulations. We will work with a flipped class-room approach for some aspects of the course.
the student understands the concept of dynamics in enzymology and can give examples of dynamic systems
the student understands the fundamentals of NMR spectroscopy and can explain the concepts of chemical shift, dipolar coupling, J-coupling, relaxation
the student understands how NMR spectroscopy can be used to study protein dynamics and can give examples and critically discuss possibilities and limitations
the student understands and can apply the theory of steady state and pre-steady state kinetics for the characterisation of enzyme-catalysed reactions in calculations and simulations
the student can explain how kinetic experiments are performed and analysed, both for steady state and stopped-flow measurements
Exercises and simulations will represent 10% of the grade. A written exam represents 90% of the grade.
Knowledge at entry:
Good understanding of protein structure (primary – quaternary structure, amino acids), basics of Michaelis Menten kinetics, concept of Boltzmann equilibrium