Flash Cooling Protein Crystals: Estimate of Cryoprotectant Concentration Using Thermal Properties

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• 作者：Binal N. Shah ; Unmesh Chinte ; Stephen J. Tomanicek ; B. Leif Hanson ; Constance A. Schall
• 刊名：Crystal Growth & Design
• 出版年：2011
• 出版时间：May 4, 2011
• 年：2011
• 卷：11
• 期：5
• 页码：1493-1501
• 全文大小：888K
• 年卷期：v.11,no.5(May 4, 2011)
• ISSN：1528-7505

文摘

X-ray diffraction from protein crystals is routinely measured at cryogenic temperatures, primarily to minimize radiation damage. Protein crystals and the surrounding mother liquor have high water content, which can lead to ice formation when samples are cooled to cryogenic temperatures. Cryoprotectants are added to the aqueous mother liquor solutions to achieve both vitreous water and to retain protein crystal integrity. Finding a minimum cryoprotectant concentration to avoid ice formation is a trial and error procedure, and valuable crystals are often lost in this exercise. To minimize crystal loss, a predictive algorithm for estimation of cryoprotectant concentration requirements for successful flash cooling was developed based on heat transfer analysis and thermal properties of cryoprotectant solutions. These thermal properties such as glass transition temperature, melting temperature, and specific heat capacity were measured and modeled for aqueous glycerol and ethylene glycol solutions, and ternary mixtures of glycerol鈭抯alt鈭抴ater. The minimum amounts of cryoprotectant required for successful flash cooling in samples of varying size were evaluated by X-ray diffraction, and a target cooling time for sample vitrification was identified. A predictive heat transfer analysis and property algorithm for cryoprotectant requirements was verified for neutron diffraction sized crystals of d-xylose isomerase and Aeropyrum pernix flap endonuclease 1.