Haymet A D, Ward L G, Harding M M, Knight C A
Department of Chemistry, University of Houston, TX 77204-5641, USA.
FEBS Lett. 1998 Jul 3;430(3):301-6. doi: 10.1016/s0014-5793(98)00652-8.
Three mutant polypeptides of the type I 37-residue winter flounder 'antifreeze' protein have been synthesized. All four threonine residues in the native peptide were been mutated to serine, valine and glycine respectively and two additional salt bridges were incorporated into the sequences in order to improve aqueous solubility. The peptides were analyzed by nanoliter osmometry, the 'ice hemisphere' test, the 'crystal habit' test, measurement of ice growth hysteresis and CD spectroscopy. While the valine and serine mutants retain the alpha-helical structure, only the valine mutant retains 'antifreeze' activity similar to that of the native protein. These data show that the threonine hydroxyl groups do not play a crucial role in the accumulation of the native 'antifreeze' protein at the ice/water interface and the inhibition of ice growth below the equilibrium melting temperature.
已合成了三种I型37个残基的冬鲽“抗冻”蛋白的突变多肽。天然肽中的所有四个苏氨酸残基分别被突变为丝氨酸、缬氨酸和甘氨酸，并且在序列中引入了另外两个盐桥以提高水溶性。通过纳升渗透压测定法、“冰半球”试验、“晶体习性”试验、冰生长滞后测量和圆二色光谱法对这些肽进行了分析。虽然缬氨酸和丝氨酸突变体保留了α-螺旋结构，但只有缬氨酸突变体保留了与天然蛋白相似的“抗冻”活性。这些数据表明，苏氨酸羟基在天然“抗冻”蛋白在冰/水界面的积累以及在平衡熔点温度以下抑制冰生长方面并不起关键作用。
