Brownlow S, Morais Cabral J H, Cooper R, Flower D R, Yewdall S J, Polikarpov I, North A C, Sawyer L
Leeds Centre for Molecular Recognition in Biological Systems, Department of Biochemistry and Molecular Biology, The University of Leeds, Leeds, LS2 9JT, UK.
Structure. 1997 Apr 15;5(4):481-95. doi: 10.1016/s0969-2126(97)00205-0.
beta-Lactoglobulin (beta-Lg) is the major whey protein in the milk of ruminants and many other mammals. Its function is not known, but it undergoes at least two pH-dependent conformational changes which may be important. Bovine beta-Lg crystallizes in several different lattices, and medium-resolution structures of orthorhombic lattice Y and trigonal lattice Z have been published. Triclinic lattice X and lattice Z crystals grow at pH values either side of the pH at which one of the pH-induced conformational changes occurs. A full understanding of the structure is needed to help explain both the conformational changes and the different denaturation behaviour of the genetic variants.
We have redetermined the structure of beta-Lg lattice Z at 3.0 A resolution by multiple isomorphous replacement and have partially refined it (R factor = 24.8%). Using the dimer from this lattice Z structure as a search model, the triclinic crystal form grown at pH 6.5 (lattice X) has been solved by molecular replacement. Refinement of lattice X at 1.8 A resolution gave an R factor of 18.1%. The structure we have determined differs from previously published structures in several ways.
Incorrect threading of the sequence in the published structures of beta-Lg affects four of the nine beta strands. The basic lipocalin fold of the polypeptide chain is unchanged, however. The relative orientation of the monomers in the beta-Lg dimer differs in the two lattices. On raising the pH, there is a rotation of approximately 5 degrees, which breaks a number of intersubunit hydrogen bonds. It is not yet clear, however, why the stability of the structure should depend so heavily upon the external loop around residue 64 or the beta strand with the free thiol, each of which shows genetic variation.
β-乳球蛋白（β-Lg）是反刍动物及许多其他哺乳动物乳汁中的主要乳清蛋白。其功能尚不清楚，但它会经历至少两种pH依赖性构象变化，这可能很重要。牛β-Lg能在几种不同的晶格中结晶，正交晶格Y和三角晶格Z的中等分辨率结构已发表。三斜晶格X和晶格Z晶体在pH诱导的构象变化之一发生的pH值两侧生长。需要对结构有全面的了解，以帮助解释构象变化和遗传变体的不同变性行为。
我们通过多重同晶置换以3.0 Å分辨率重新测定了β-Lg晶格Z的结构，并对其进行了部分精修（R因子 = 24.8%）。使用该晶格Z结构中的二聚体作为搜索模型，通过分子置换解析了在pH 6.5时生长的三斜晶型（晶格X）。以1.8 Å分辨率对晶格X进行精修，得到的R因子为18.1%。我们确定的结构在几个方面与先前发表的结构不同。
β-Lg已发表结构中序列的错误穿线影响了九条β链中的四条。然而，多肽链的基本脂质运载蛋白折叠未变。β-Lg二聚体中单体的相对取向在两种晶格中有所不同。提高pH值时，会有大约5度的旋转，这会破坏一些亚基间的氢键。然而，目前尚不清楚为什么结构的稳定性会如此严重地依赖于残基64周围的外部环或带有游离巯基的β链，这两者都表现出遗传变异。
