Toolbox: protein analysis techniques

Duration

20h Th, 20h AUTR

Number of credits

Lecturer

Christian Damblon, Franck Dequiedt, Mireille Dumoulin, André Matagne, N..., Damien Sluysmans, Marylène Vandevenne

Coordinator

André Matagne

Language(s) of instruction

French language

Organisation and examination

Teaching in the first semester, review in January

Schedule

Schedule online

Units courses prerequisite and corequisite

Prerequisite or corequisite units are presented within each program

Learning unit contents

1. Aim of the module

Various analytical methods used for protein characterisation are presented. Thus, spectroscopic methods are UV-visible absorption, linear and circular dichroism, fluorescence, vibrational spectroscopy (i.e. infrared and Raman), surface plasmon resonance (SPR) and biolayer interferometry (BLI). Three 'single molecule' approaches (i.e. FRET, AFM and optical tweezers) are also studied, as well as methods for measuring interactions between molecules in real time. Classical thermodynamic approaches (i.e. DSC, ITC, DSF) are presented, as well as methods specific to the study of in vivo interactions. For each approach, the principles, technical aspects, and applications focusing on proteins will be developed, especially during the article presentation sessions and the practical work (demonstrations) that will take place in the afternoons. The selected articles focus mainly on the stability and folding (i.e. the process of acquiring the 3D structure) of proteins. A few models (e.g. beta-lactamases, single-domain antibody fragments, lysozymes) are analysed in detail, based on theoretical knowledge and data found in the literature.

 

2. Table of contents of theoretical courses

2.1. Optical spectroscopy applied to the study of biological macromolecules (UV-Vis absorption; infrared and Raman; linear and circular dichroism; fluorescence). 9h; Christian Damblon

2.2. Static and dynamic light scattering (SLS and DLS, respectively) for the analysis of biological macromolecules. 2h; Marylène Vandevenne

 2.3. Thermodynamic methods for the analysis of biological macromolecules and the study of their interactions: a) isothermal titration calorimetry (ITC); b) differential scanning calorimetry (DSC); c) differential scanning fluorimetry (DSF). 2h; Marylène Vandevenne

2.4. Real-time biosensing methods based on e.g. SPR, BLI, QCM. 2h; Mireille Dumoulin

2.5. Single molecule approaches (AFM, optical tweezers and FRET). 3h; Damien Sluysmans & Christian Damblon

2.6. In vivo interactions (two-hybrid screening, protein complementation assay, etc.). 2h; Frank Dequiedt

 

3. Tables of contents of practical courses and tutorials

The use of biophysical methods to study protein folding and stability is illustrated on the basis of data found in the litterature (André Matagne).

Demonstrations are organised using the instruments available at the CIP (André Matagne, Marylène Vandevenne, Romain Malempré).

Learning outcomes of the learning unit

Students will gain an overview of the main analytical tools available to characterise proteins at the molecular level. Particular attention will be paid to the biophysical methods available for characterising protein stability and folding, as well as for studying the conformational dynamics of biological macromolecules in general. Students should be able to select the most relevant approaches for characterising a protein sample. These methods are important both for basic biochemical characterisation, particularly with a view to more advanced structural studies, and for quality control in the preparation of samples for e.g. biomedical applications.

Prerequisite knowledge and skills

Basic chemistry and physics. Fundamental knowledge of the structure of biological macromolecules

Planned learning activities and teaching methods

Lectures are given with the help of powerpoint presentations, blackboard and chalk

Mode of delivery (face to face, distance learning, hybrid learning)

Face-to-face course


Additional information:

Face-to-face only

Recommended or required readings

Powerpoint presentations and/or lecture notes will be available at the end of the course at the latest. These materials are intended to assist students in their study; not all of them are absolutely essential for acquiring the skills required in this course and their adoption by students is not compulsory. Scientific articles will be given as they are analyzed.

Reference textbooks :

- Biophysical Chemistry, Cantor and Schimmel

- Principles of Physical Chemistry, Van Holde et al.

- Biological Spectroscopy, Campbell and Dwek

- Physical Biochemistry, Sheehan

- The Physical and Chemical Basis of Molecular Biology, Creighton

- The Biophysical Chemistry of Nucleic Acids and Proteins, Creighton

 

Exam(s) in session

Any session

- In-person

oral exam


Additional information:

An oral exam will be organised in the presence of the module coordinator (André Matagne) and Professor Christian Damblon. The presence of the other lectures (Frank Dequiedt, Mireille Dumoulin, Damien Slysmans, Marylène Vandevenne) is optional. The evaluation will cover the whole subject, in the form of an integrated approach which encourages reflection and invites the student to take into consideration all the tools likely to enable the complete characterisation of a protein (or possibly any other biological macromolecule). The discussion will focus on the series of articles seen in class and others recommended for reading by teachers.

Work placement(s)

Non applicable

Organisational remarks and main changes to the course

Lectures will be organized following the schedule known by the students. The professor will
inform the students in advance of any possible modification to this

Contacts

André Matagne, PhD, Professeur Ordinaire, Enzymologie et Repliement des Protéines, Centre
d'Ingénierie des Protéines, UR InBioS, Département des Sciences de la Vie, Institut de Chimie B6a, (bureau 1/10a), Quartier Agora, Allée du 6 Août, 13, Université de Liège, B4000 Liège (Sart-Tilman), Tel.: +32 (0)4 3663419, Email: amatagne@ulg.ac.be (best option)

Association of one or more MOOCs

There is no MOOC associated with this course.


Additional information:

Non applicable