OPPF-MPL High-throughput Protein Production and Crystallization

Training
Date: 30-Mar-2011 1:00 CEST to -

Contact:

Aim

The aim of this course is to review the state-of-the-art in HTP structural biology with an emphasis on methods to study complex and "difficult" targets, including membrane proteins.

Participants

The course is aimed at Post-doctoral researchers and PhD students with at least one year's relevant experience in structural biology. The course will be limited to 12 participants and priority will be given to applicants from UK Academic Institutions.

Course Content

The course will be structured so as to interleave lectures and practical sessions. The core of the course will be followed by all participants, although towards the end of the course there will be the possibility of selection a topic from a number of specialist modules, such as thermafluor studies of protein stability and mass spectrometry as a routine tool for quality assessment, detergent screening for membrane protein purification. The core activity will cover the process of HTP protein production from target selection (bioinformatics) through parallel cloning (96 well format), parallel protein expression screening (both E. coli and eukaryotic cells), protein expression scale-up and purification and crystallization. Participants will be able (via a pro-active organization prior to the course) to bring their own targets, which they will be able to enter into the process at any of these points, for example, cDNAs for cloning and expression and purified proteins for crystallization studies. Where possible, work will follow the flow of actual experiments, although to cover the full range of activities some aspects will be run off-line (for instance bioinformatics and target selection will be covered whilst the cloning experiments are on-going).

The course will therefore cover the following:-

  1. Target selection / LIMS (lecture + practical)
  2. HTP cloning + expression screening (lecture + practicals)
  3. HTP baculovirus/mammalian expression construction (lecture + practical)
  4. HTP protein purification and characterization (lecture + practical)
  5. HTP protein crystallization (lectures + practical)