Crystallization and preliminary X-ray crystallographic analysis of an ice-binding protein (FfIBP) from Flavobacterium frigoris PS1

Abstract

Ice growth in a cold environment is fatal for polar organisms, not only because of the physical destruction of inner cell organelles but also because of the resulting chemical damage due to processes such as osmotic shock. The properties of ice-binding proteins (IBPs), which include antifreeze proteins (AFPs), have been characterized, and IBPs exhibit the ability to inhibit ice growth by binding to specific ice planes and lowering the freezing point. Our lab has recently succeeded in over-expressing an ice-binding protein (FfIBP) from the Gram-negative bacteria Flavobacterium frigoris PS1, which was isolated from the Antarctic. Interestingly, the thermal hysteresis activity of FfIBP was approximately 2.5°C at 50 ？M, which is 10 times higher than that of the moderately active IBP from Arctic yeast (LeIBP). Although FfIBP closely resembles LeIBP in its amino acid sequence, the antifreeze activity of FfIBP appears to be much greater than that of LeIBP. In an effort to understand the reason for this difference, we attempted to solve the crystal structure of FfIBP. Herein, the crystallization methods and X-ray diffraction data for FfIBP are reported. FfIBP was crystallized using the hanging drop vapor diffusion method with 0.1 M sodium acetate at pH 4.4 and 3 M sodium chloride as a precipitant. A complete diffraction data set was collected at a resolution of 2.9 ？. The crystal belos of ice-binding proteins (IBPs), which include antifreeze proteins (AFPs), have been characterized, and IBPs exhibit the ability to inhibit ice growth by binding to specific ice planes and lowering the freezing point. Our lab has recently succeeded in over-expressing an ice-binding protein (FfIBP) from the Gram-negative bacteria Flavobacterium frigoris PS1, which was isolated from the Antarctic. Interestingly, the thermal hysteresis activity of FfIBP was approximately 2.5°C at 50 ？M, which is 10 times higher than that of the moderately active IBP from Arctic yeast (LeIBP)