Methods in Biochemical Investigation

Teaching Unit of Structural Analysis (3 CFU)
This teaching module is focused on the 3D structure determination of proteins by using biocrystallographic techniques. The method for structure determination through X-ray diffraction will be analyzed, with emphasis on the experimental techniques for protein crystal growth, and on the basic principles of X-ray diffraction: X-ray sources, synchrotron light, atomic scattering factors, structure factors, the "phase problem", phasing by molecular replacement and heavy atoms. Protein model refinement and validation techniques will be analyzed, together with the basic concepts and contents of the Protein Data Bank.

Teaching unit of Form Advanced Enzymology (3 CFU)
General aim of the teaching unit is to increase students' understanding of enzyme properties, building on the bases provided by the first-level degree in Biological Sciences. The following topics will be covered: Enzyme reaction mechanisms and related rate equations under steady-state conditions. One- and two-substrate reactions. Reversible enzyme reactions. Techniques for the study of steady-state phase of enzyme reactions. Enzyme assays. Measurement of enzyme kinetic parameters. Enzyme inhibition. Experimental design and interpretation of results in the kinetic study of enzyme reactions. Protein engineering for the study of structure-function relationships in enzymes at the molecular level.


Reference Material
Teaching Unit of Structural Analysis:
- Principles of protein X-ray crystallography (2nd edition). J. Drenth (Springer)
- Crystallography made crystal clear. A guide for users of macromolecular models. By Gale Rhodes (Elsevier)

Teaching unit of Advanced Enzymology:
Students can refer to their text of Biochemistry for the general aspects of the structure of enzymes and enzymatic catalysis. Further useful insights can be found in:
- N. Price and L. Stevens, "Fundamentals of Enzymology - Cell and Molecular Biology of Catalytic Proteins" 3rd Ed., Oxford University Press, 1999.
- A. Fersht, "Structure and Mechanism in Protein Science. A Guide to Enzyme Catalysis and Protein Folding ", WH Freeman & Co., 1999


Prerequisites and examination procedures
Written examination with questions with open answers for the Structural Biology part, written or oral (your choice) with questions with open answers and exercises for the Advanced Enzymology part.


Teaching Methods
Traditional. Lectures supported by projected material.
Attendance: highly recommended.


Language of instruction
English


Program information
Copies of the slides projected in the classroom as well as other materials will be made available through the course website on the ARIEL platform of the University of Milano (http://ariel.ctu.unimi.it). By no means this material replaces the lectures or a textbook. The material is made available only to registered students of the Degree Course in Biological Sciences and should not be distributed to others.


WEB pages
http://mnardinimib.ariel.ctu.unimi.it/v3/home/Default.aspx