As part of a structure-based drug-discovery effort at RAPID (Rational Approaches to Pathogen Inhibitor Discovery), IspD from Mycobacterium tuberculosis was targeted. Two years of effort finally resulted in three crystal structures of mycobacterial IspDs, in complex with substrates (1). Obtaining the crystals proved to be challenging.
In this talk, I will present the approaches that worked, as well as the ones that did not. At RAPID, we employ a minimalist approach to screening chemical space. How well did this strategy serve in the IspD project? The choice of construct, screening kits, buffer, protein concentration, seeding, and crystallization strategy will also be discussed. Our experiences in this project may be of relevance to other small academic laboratories with limited automation for screening crystallization.
(1) Björkelid C, Bergfors T, Henriksson LM , Stern AL, Unge, T Mowbray SL and Jones TA. Structural and functional studies of mycobacterial IspD enzymes. Acta Cryst 2011 D67, 403-414.