Lefèvre T, Subirade M
Université Laval, Faculté des sciences de l'agriculture et de l'alimentation, Centre de recherches en Sciences et Technologie du Lait (STELA), Pavillon Paul Comtois, Sainte-Foy, Québec, Canada G1K 7P4.
Biopolymers. 2000 Dec;54(7):578-86. doi: 10.1002/1097-0282(200012)54:7<578::AID-BIP100>3.0.CO;2-2.
In order to reveal at a molecular level differences between fine-stranded and particulate gels, we present an Fourier transform infrared spectroscopic study of the thermal behavior of beta-lactoglobulin (beta-lg) in salt-free D(2)O solutions and low ionic strength at different pDs. Differences are found in the denaturation mechanism, in the unfolded state of the protein, in the aggregate formation, and in the strength of the intermolecular interactions. For fine-stranded gels (pD 2.8 and 7.8), heating induces the dissociation of the dimers into monomers. The protein undergoes extensive structural modifications before aggregation begins. Aggregation is characterized by the appearance of a new band attributed to intermolecular beta-sheets which is located in the 1613-1619 cm(-1) range. For particulate gels (pD 4.4 and 5.4), the protein structure is almost preserved up to 75-80 degrees C with no splitting of the dimers. The band characteristic of aggregation originates from the component initially located at 1623 cm(-1), suggesting that at the beginning of aggregation, globular beta-lg in the dimeric form associate to constitute oligomers with higher molecular mass. Aggregation may result in the association of globular slightly denatured dimers, leading to the formation of spherical particles rather than linear strands. The aggregation band is always located in the 1620-1623 cm(-1) range for particulate gels showing that hydrogen bonds are weaker for these aggregates than for fine-stranded ones. This has been related to a more extensive protein unfolding for fine-stranded gels that allows a closer alignment of the polypeptide chains, and then to the formation of much stronger hydrogen bonds. Small differences are also found in protein organization and in intermolecular hydrogen bond strength vs pD within the same type of gel. Protein conformation and protein-protein interactions in the gel state may be responsible of the specific macroscopic properties of each gel network. A coarse representation of the different modes of gelation is described.
为了在分子水平上揭示细链凝胶和颗粒凝胶之间的差异，我们对不同pD值下无盐D₂O溶液和低离子强度条件下β-乳球蛋白（β-lg）的热行为进行了傅里叶变换红外光谱研究。在变性机制、蛋白质的未折叠状态、聚集体形成以及分子间相互作用强度方面发现了差异。对于细链凝胶（pD 2.8和7.8），加热会导致二聚体解离成单体。在聚集开始之前，蛋白质会经历广泛的结构修饰。聚集的特征是出现一个归因于分子间β-折叠的新谱带，其位于1613 - 1619 cm⁻¹范围内。对于颗粒凝胶（pD 4.4和5.4），蛋白质结构在高达75 - 80℃时几乎保持不变，二聚体没有分裂。聚集的特征谱带最初位于1623 cm⁻¹处，这表明在聚集开始时，二聚体形式的球状β-lg缔合形成更高分子量的寡聚体。聚集可能导致球状轻度变性的二聚体缔合，从而形成球形颗粒而非线性链。对于颗粒凝胶，聚集谱带始终位于1620 - 1623 cm⁻¹范围内，这表明这些聚集体中的氢键比细链凝胶中的氢键弱。这与细链凝胶中更广泛的蛋白质展开有关，这种展开允许多肽链更紧密地排列，进而形成更强的氢键。在同一类型的凝胶中，还发现了蛋白质组织以及分子间氢键强度与pD之间的微小差异。凝胶状态下的蛋白质构象和蛋白质 - 蛋白质相互作用可能是每种凝胶网络特定宏观性质的原因。描述了不同凝胶化模式的粗略表示。
