Biochemical and biophysical parameters influencing macromolecular crystallization and X-ray diffraction quality

Abstract

Póster presentado en el Congrés Internacional de Biologia de Catalunya "Global Questions on Advanced Biology : an international congress on interdisciplinary frontiers in biology", organizado por la Societat Catalana de Biologia, del 9 al 12 de julio de 2012 en Barcelona (España)One way to investigate and properly understand the function of a protein and its interaction with partners is to know its three-dimensional structure. Macromolecular crystallography is a tool that provides the three-dimensional structure at atomic level of a protein that has been previously crystallized. A protein crystal consists of a very large number of repeating units where each individual unit is known as the unit cell, with no internal crystalline symmetry and which contains the crystallized sample. In general, crystallization starts with the formation of nuclei of protein molecules in supersaturated chemical conditions. There are several techniques available for bringing a pure protein solution gradually to a supersaturated state, such as batch, microbatch, vapour diffusion by hanging or sitting drops, and seeding. Once obtained a protein crystal, a potential bottleneck is to obtain a wellordered crystal that will diffract X-rays strongly. Sometimes co-crystallization of a protein with a substrate may help the crystal quality, because the protein is structurally stabilized by the ligand, the crystal packing is more regular and this improves the X-ray diffraction pattern. In this case, a protein in complex with different substrates may result in different crystals that yield X-ray diffraction patterns of variable quality. We will present an example of crystal quality improvement of a protein/DNA complex in which we changed the design of the oligonucleotides harboring the DNA binding site, including the sequence, the length and the type of ends, blunt or cohesive. These changes modified the crystallization, as assessed by the macroscopic aspect of the crystals and the corresponding X-ray diffraction qualityPeer Reviewe