Department of Food Science, Aarhus University, Agro Food Park 48, 8200 Aarhus N, Denmark.
Department of Food Science, Aarhus University, Agro Food Park 48, 8200 Aarhus N, Denmark.
J Dairy Sci. 2022 Mar;105(3):1959-1965. doi: 10.3168/jds.2021-21172. Epub 2022 Jan 5.
Variations in the phosphorylation and glycosylation patterns of the common κ-casein (CN) variants A and B have been explored, whereas studies on variant E heterogeneity are scarce. This study reports for the first time the detailed phosphorylation and glycosylation pattern of the κ-CN variant E in comparison with variants A and B. Individual cow milk samples representing κ-CN genotype EE (n = 12) were obtained from Swedish Red cows, and the natural posttranslational modifications of its κ-CN were identified and quantified by liquid chromatography-electrospray mass spectrometry. In total, 12 unique isoform masses of κ-CN variant E were identified. In comparison, AA and BB milk consisted of 14 and 17 unique isoform masses, respectively. The most abundant κ-CN E isoform detected in the EE milk was the monophosphorylated, unglycosylated [1P 0G, ∼70%; where P indicates phosphorylation from single to triple phosphorylation (1-3P), and G indicates glycosylation from single to triple glycosylation (1-3G)] form, followed by diphosphorylated, unglycosylated (2P 0G, ∼12%) form, resembling known patterns from variants A and B. However, a clear distinction was the presence of the rare triphosphorylated, nonglycosylated (3P 0G, ∼0.05%) κ-CN isoform in the EE milk. All isoforms detected in variant E were phosphorylated, giving a phosphorylation degree of 100%. This is comparable with the phosphorylation degree of variants A and B, being also almost 100%, though with very small amounts of nonphosphorylated, glycosylated isoforms detected. The glycosylation degree of variant E was found to be around 17%, a bit higher than observed for variant B (around 14%), and higher than variant A (around 7%). Among glycosylation, the glycan e was the most common type identified for all 3 variants, followed by c/d (straight and branched chain trisaccharides, respectively), and b. In contrast to κ-CN variants A and B, no glycan of type a was found in variant E. Taken together, this study shows that the posttranslational modification pattern of variant E resembles that of known variants to a large extent, but with subtle differences.
κ-酪蛋白（CN）变体 A 和 B 的磷酸化和糖基化模式已经得到了广泛研究，而关于变体 E 异质性的研究则相对较少。本研究首次报道了 κ-CN 变体 E 与变体 A 和 B 相比的详细磷酸化和糖基化模式。从瑞典红牛中获得了代表 κ-CN 基因型 EE（n=12）的个体牛奶样本，并通过液相色谱-电喷雾质谱法对其 κ-CN 的天然翻译后修饰进行了鉴定和定量。总共鉴定出 12 种独特的 κ-CN 变体 E 同工型质量。相比之下，AA 和 BB 牛奶分别由 14 和 17 种独特的同工型质量组成。在 EE 牛奶中检测到的最丰富的 κ-CN E 同工型是单磷酸化、未糖基化的[1P0G，约 70%；其中 P 表示从单磷酸化到三磷酸化（1-3P）的磷酸化，G 表示从单糖基化到三糖基化（1-3G）的糖基化]形式，其次是双磷酸化、未糖基化（2P0G，约 12%）形式，类似于已知的变体 A 和 B 的模式。然而，一个明显的区别是在 EE 牛奶中存在罕见的三磷酸化、非糖基化（3P0G，约 0.05%）κ-CN 同工型。在变体 E 中检测到的所有同工型都被磷酸化，磷酸化程度为 100%。这与变体 A 和 B 的磷酸化程度相当，也接近 100%，尽管检测到的非磷酸化、糖基化同工型非常少。变体 E 的糖基化程度约为 17%，略高于变体 B（约 14%），高于变体 A（约 7%）。在糖基化中，e 型聚糖是所有 3 种变体中最常见的类型，其次是 c/d（直链和支链三糖，分别）和 b。与变体 A 和 B 不同，变体 E 中未发现 a 型聚糖。总的来说，本研究表明变体 E 的翻译后修饰模式在很大程度上与已知变体相似，但存在细微差异。
