Harding M M, Ward L G, Haymet A D
School of Chemistry, University of Sydney, NSW 2006, Australia.
Eur J Biochem. 1999 Sep;264(3):653-65. doi: 10.1046/j.1432-1327.1999.00617.x.
The type I 'antifreeze' proteins, found in the body fluids of fish inhabiting polar oceans, are alanine-rich alpha-helical proteins that are able to inhibit the growth of ice. Within this class there are two distinct subclasses of proteins: those related to the winter flounder sequence HPLC6 and which contain 11-residue repeat units commencing with threonine; and those from the sculpins that are unique in the N-terminal region that contains established helix breakers and lacks the 11-residue repeat structure present in the rest of the protein. Although 14 type I proteins have been isolated, almost all research has focused on HPLC6, the 37-residue protein from the winter flounder Pseudopleuronectes americanus. This protein modifies both the rate and shape (or 'habit') of ice crystal growth, displays hysteresis and accumulates specifically at the {2 0 2; 1} ice plane. Until very recently, all models to explain the mechanism for this specific interaction have relied on the interaction of the four threonine hydroxyls, which are spaced equally apart on one face of the helix, with the ice lattice. In contrast, proteins belonging to the sculpin family accumulate specifically at the {2 1; 1; 0} plane. The molecular origin of this difference in specificity between the flounder and sculpin proteins is not understood. This review will summarize the structure-activity and molecular modelling and dynamics studies on HPLC6, with an emphasis on recent studies in which the threonine residues have been mutated. These studies have identified important hydrophobic contributions to the ice growth inhibition mechanism. Some 50 mutants of HPLC6 have been reported and the data is consistent with the following requirements for ice growth inhibition: (a) a minimum length of approx. 25 residues; (b) an alanine-rich sequence in order to induce a highly helical conformation; (c) a hydrophobic face; (d) a number of charged/polar residues which are involved in solubility and/or interaction with the ice surface. The emerging picture, that requires further dynamics studies including accurate modelling of the ice/water interface, suggests that a hydrophobic interaction between the surface of the protein and ice is the key to explaining accumulation at specific ice planes, and thus the molecular level mechanism for ice growth inhibition.
I型“抗冻”蛋白存在于栖息在极地海洋鱼类的体液中，是富含丙氨酸的α螺旋蛋白，能够抑制冰的生长。在这一类蛋白中，有两个不同的亚类：一类与冬比目鱼序列HPLC6相关，含有以苏氨酸开头的11个残基重复单元；另一类来自杜父鱼，其N端区域独特，含有已确定的螺旋破坏子，且缺乏该蛋白其余部分所具有的11个残基重复结构。虽然已经分离出14种I型蛋白，但几乎所有研究都集中在HPLC6上，它是来自美洲拟庸鲽的一种37个残基的蛋白。这种蛋白既能改变冰晶生长的速率，也能改变其形状（或“习性”），具有滞后现象，并特异性地聚集在{2 0 2; 1}冰面上。直到最近，所有解释这种特异性相互作用机制的模型都依赖于螺旋一侧等间距排列的四个苏氨酸羟基与冰晶格的相互作用。相比之下，杜父鱼家族的蛋白特异性地聚集在{2 1; 1; 0}平面上。比目鱼和杜父鱼蛋白在特异性上的这种差异的分子起源尚不清楚。这篇综述将总结关于HPLC6的结构活性、分子建模和动力学研究，重点是最近对苏氨酸残基进行突变的研究。这些研究已经确定了对冰生长抑制机制的重要疏水贡献。已经报道了大约50种HPLC6的突变体，数据与冰生长抑制的以下要求一致：（a）最小长度约为25个残基；（b）富含丙氨酸的序列，以诱导高度螺旋构象；（c）一个疏水表面；（d）一些参与溶解性和/或与冰表面相互作用的带电/极性残基。新出现的情况，需要进一步的动力学研究，包括对冰/水界面的精确建模，表明蛋白表面与冰之间的疏水相互作用是解释在特定冰面上聚集的关键，从而也是冰生长抑制的分子水平机制。
