07-第三讲 蛋白质结晶-2011课件.pptVIP

Protein Crystallization ;Crystal Structural Strategies;Outlines;Frequently Asked Questions;Lattices: primitive or non-primitive(2) (Bravais Lattices: 14) Physical/chemical properties: Covalent Crystals, Metallic Crystals, Ionic Crystals, Molecular Crystals Q: How do crystals form and how do they grow? Phase diagram: unsaturated, supersaturated, nucleation(unasisted, asisted) ;Q: Why do different crystals have different shapes and sizes? The internal symmetry of the crystal The relative growth rates along the various directions in the of the crystal Q: What do you consider a perfect crystal? A perfect crystal is one which has a single lattice (i.e. not a twinned crystal) and is completely regular, free of defects and dislocations.;THEORY: phase diagram;An energy barrier to crystallization ;The probability of nucleation increases with increasing supersaturation The more supersaturated the protein solution, the greater the likelihood that a critical nucleus will form the smaller the nucleus needed to induce crystal formation This can be represented on a phase diagram by dividing the supersaturated zone into regions of increasing probability of nucleation and precipitation. ;PROTEIN: know your protein well;Is it fresh? Proteins break down with time and the mixture becomes heterogeneous Always try to set up your trials as soon as possible, preferably the same day as the last step of the purification Is it monodispersed? Monodispersity means that the protein exists in solution as a single oligomeric species. It means the protein is free of non-specific oligomers and aggregates. Use a size-exclusion column as the last step in the purification. Use the DLS to see if your protein is monodispersed. ;Does your protein need to be kept reduced? Does your protein need the addition of something (eg salt) to stay in solution? Is your protein stable at room temperature? Does your protein break down rapidly? Has anything similar been crystallized before? Chec