Institute for Pharmaceutical and Medicinal Chemistry, University of Münster, PharmaCampus, Correnstr. 48, 48149 Münster, Germany.
Department of Rheumatology and Hiller Research Unit Rheumatology, Medical Faculty, Heinrich-Heine-University Düsseldorf, Moorenstr. 5, 40225 Düsseldorf, Germany.
Int J Mol Sci. 2024 Feb 1;25(3):1743. doi: 10.3390/ijms25031743.
Breast-milk α-casein is a Toll-like receptor 4 (TLR4) agonist, whereas phosphorylated α-casein does not bind TLR4. The objective of this study was to analyse the structural requirements for these effects. In silico analysis of α-casein indicated high α-helical content with coiled-coil characteristics. This was confirmed by CD-spectroscopy, showing the α-helical conformation to be stable between pH 2 and 7.4. After in vitro phosphorylation, the α-helical content was significantly reduced, similar to what it was after incubation at 80 °C. This conformation showed no in vitro induction of IL-8 secretion via TLR4. A synthetic peptide corresponding to V-E of α-casein induced an IL-8 secretion of 0.95 ng/mL via TLR4. Our results indicate that α-casein appears in two distinct conformations, an α-helical TLR4-agonistic and a less α-helical TLR4 non-agonistic conformation induced by phosphorylation. This is to indicate that the immunomodulatory role of α-casein, as described before, could be regulated by conformational changes induced by phosphorylation.
母乳α-酪蛋白是 Toll 样受体 4（TLR4）激动剂，而磷酸化的α-酪蛋白不与 TLR4 结合。本研究旨在分析这些作用的结构要求。α-酪蛋白的计算机分析表明其具有高α-螺旋含量和卷曲螺旋特征。这通过 CD 光谱学得到了证实，表明在 pH 2 至 7.4 之间，α-螺旋构象稳定。体外磷酸化后，α-螺旋含量显著降低，与 80°C 孵育后的结果相似。这种构象在体外没有通过 TLR4 诱导 IL-8 分泌。与 α-酪蛋白的 V-E 相对应的合成肽通过 TLR4 诱导 IL-8 分泌 0.95ng/mL。我们的结果表明，α-酪蛋白呈现出两种不同的构象，一种是 TLR4 激动性的α-螺旋构象，另一种是由磷酸化诱导的较少α-螺旋 TLR4 非激动性构象。这表明，如前所述，α-酪蛋白的免疫调节作用可能受到磷酸化诱导的构象变化的调节。
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