Cryo Electron Microscopy in Structural Biology - Einzelansicht

Content:

Structual biology is the study of how biological molecules are built. Only if we know their structure, we can deduce important information on their function, their assembly and their ability to interact. Cryo electron microscopy has revolutionised structural biology in the recent years (https://www.nature.com/articles/d41586-020-00341-9)

The course covers theory and advanced image analysis techniques in transmission electron microscopy (TEM). In the lecture the following topics are explained:
- buildup of a TEM, electron guns, holders and detectors
- theory of diffraction, image formation, how electrons interact with material; contrast transfer function
- theory behind high resolution cryo TEM
- sample preparation and image acquisition in (single particle) TEM
- 2D and 3D image analysis
- point group symmetries
- refinement and validation of 3D reconstructions
- visualisation and interpretation of TEM results

During the seminar the students will analyse current TEM results from subject-specific English literature, summarize the key aspects of the work and discuss the research outcome critically.

The practical course will cover hands-on practical work on the transmission electron microscope as well as single-particle image processing of a high-resolution cryo EM data set. The TEM work will include
- negative staining of a protein complex
- loading of the grid into the NS holder
- setting up a negative stain data collection in the data acquisition software
- freezing of a cryo EM grid
- demonstration of the entire cryo EM workflow

The image processing will be undertaken on the teaching server of the IBB using the computer pools in building 25 (Campus Golm). Students will learn how to work with the linux based server system and the EM processing software Relion. Students will learn how to assess the quality of the data and how to get from individual images to a high-resolution 3D model of the protein complex.

Learning outcomes:
1. Subject-specific competences:
At the end of the course the students will have learned and understood the underlying theory in transmission electron microscopy and single particle image analysis. They will be able to identify suitable TEM applications to a biological question and will have gained insight into the current state of the art in cryo electron microscopy. They will also have analysed the structure and function of diverse biological complexes.
2. Methods-specific competences:
The students learn to interpret, analyse and present results derived from subject-specific, English literature. They will also learn how to prepare a TEM sample, how to operate a TEM, how to analyse TEM images and how to interpret TEM data.
3. Action competence:
The students present and defend scientific work in a public seminar using suitable presentation media. The students learn to work in a team, give constructive feedback and assess each other in a public seminar. When participating in the practical course students will learn to collect and assess scientific data. Furthermore they will summarize experimental data in a detailed protocol and evaluate their results.

This course can be taken as a 6CP, 8CP or 11CP module:
6CP: Lecture and seminar
8CP: Lecture, seminar and 2 week practical course
11CP: Lecture seminar and 6 week practical course

Moodle Page: https://moodle2.uni-potsdam.de/course/view.php?id=18544

The course will go through the online resources on the Caltech webpage:

http://cryo-em-course.caltech.edu/videos

interesting review articles:

https://www.sciencedirect.com/science/article/pii/S0304416517302374

https://dasher.wustl.edu/bio5357/readings/molcell-58-677-15.pdf

interesting short youtube videos:

https://www.youtube.com/watch?v=BJKkC0W-6Qk

https://www.youtube.com/watch?v=026rzTXb1zw&t=16s

https://www.youtube.com/watch?v=Qq8DO-4BnIY