The process of forming a protein crystal from a protein solution. These crystals are then used to study the protein’s three-dimensional structure using techniques like X-ray crystallography.
Before proteins are in the crystalline state, they are typically in a dissolved state within a solution. In this state, the protein molecules are dispersed throughout the solvent (often water or a buffer solution) and are free to move around, though they might still maintain their native folded structure. The goal of protein crystallization is to transition these dispersed protein molecules from the solution into an ordered, solid lattice arrangement, forming a crystal. This transition is facilitated by finding the right conditions (e.g., concentration, pH, temperature, and presence of certain salts or other molecules) that encourage the protein molecules to come together and form this ordered structure.
Protein crystallization is a pivotal technique in molecular biology, enabling scientists to visualize proteins at atomic resolution. This detailed view offers profound insights into various aspects of biology and medicine including
The OT-2 is a bench-top liquid handler designed to be accessible and flexible enough to automate many common applications.
Overview: Vapor diffusion is a widely used method for protein crystallization that relies on the slow concentration of both the protein and the precipitant. As water evaporates from the protein solution to a reservoir solution, the protein and precipitant concentrations increase. Once the solution reaches supersaturation, crystals can form.
Setup: In both the hanging drop and sitting drop variations, a drop containing a mixture of protein solution and precipitant is placed in proximity to a reservoir containing a higher concentration of the precipitant. In the hanging drop method, the drop is placed on an inverted cover slip sealed over the reservoir. In the sitting drop method, the drop sits on a platform within a sealed well, above the reservoir.
Mechanism: Over time, water from the drop vaporizes and diffuses to the reservoir. This evaporation concentrates the protein and precipitant in the drop, promoting crystallization under favorable conditions.
Overview: In this method, the protein and precipitant are mixed together directly, and the system is left undisturbed to allow crystals to form.
Setup: A solution of the protein is mixed with a solution of the precipitant in a single container.
Mechanism: As conditions within the mixture become favorable (often due to slow evaporation or due to the specific interactions between the protein molecules and the precipitant), protein molecules start to come together and arrange in an ordered manner, forming crystals.
Overview: This is a variation of the batch method but is designed for very small volumes and to prevent evaporation.
Setup: Small volumes of protein and precipitant are mixed together in a well. This mixture is then covered with a layer of oil (like paraffin or silicone oil) to prevent evaporation.
Mechanism: The oil layer ensures that the conditions within the drop remain stable over time. Without the influence of evaporation, the protein and precipitant can interact in a controlled manner, leading to crystallization under the right conditions.
Opentrons helps you automate Protein Crystallization with open-source protocols for the OT-2 and Opentrons Flex