The aim of the course is to provide students with a thorough overview of the most important and recent methods applied to structural biology, focusing in particular on the scientific objectives that these techniques can help to achieve. After following this course, the students will develop a better capability to analyze the correlation between structure and function of proteins, and a better understanding of the experimental strategies applied in this research field. The course is ideally linked to those dealing with protein engineering and structural bioinformatics.
The course is focused on the 3D structure determination and analysis of proteins by using biocrystallographic and bioinformatic techniques. Experimental techniques for protein crystal growth will be described in details, with application to soluble and membrane proteins. X-ray diffraction on protein crystals and on solution samples (SAXS) will be treated extensively (X-ray sources, synchrotron light, atomic scattering factors, structure factors, the "phase problem", phasing by molecular replacement and heavy atoms, crystallographic refinement). Protein model validation techniques will be analyzed, together with the main features of the Protein Data Bank. In addition, the theoretical basis and the practical applications of single-molecule cryo-EM techniques will be treated. The last part of the course will have a more practical approach. Students will solve by molecular replacement, refine, and analyze the 3D structure of a protein. During this practical experience they will learn how to apply techniques and software for protein sequence alignment, secondary structure prediction, molecular modelling, crystallographic refinement and validation.
Informazioni sul programma
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 Molecular Biotechnology and Bioinformatics and should not be distributed to others.
Prerequisiti e modalità di esame
The evaluation of the student's performance is based on a written examination with 4-5 open-answer questions spanning all topics covered in the course. A written examination allows students to show their ability to describe and critically comment (by using diagrams, graphs, equations) the theoretical bases and the practical applications of the structural biology methods learned during the course. No exercises are present in the examination test. Examples of the examination test will be discussed during classes and made available to students.
Teaching mode: classroom lectures supported by projected material, visit to the biocrystallography lab and practical sessions in a cyber lab. Attendance: highly recommended.
Materiale didattico e bibliografia
Principles of protein X-ray crystallography (2nd edition). J. Drenth (Springer) Crystallization of nucleic acids and proteins: a practical approach. Edited by A. Ducruix and R. Giegé (Oxford University press) Crystallography made crystal clear. A guide for users of macromolecular models. By Gale Rhodes (Elsevier)