Lima Maria, Moloney Catherine, Ames Jennifer M
Human Nutrition and Health Group, School of Biological Sciences, Queen's University Belfast, David Keir Building, Stranmillis Road, Belfast BT9 5AG, Northern Ireland, UK.
Amino Acids. 2009 Mar;36(3):475-81. doi: 10.1007/s00726-008-0105-y. Epub 2008 May 31.
Modification of protein by carbonyl compounds under in vitro physiological conditions is site-directed. There are few reports of the site specificity of glycation of proteins using heating conditions of relevance to food processing. The aim of this study was to determine the site specificity of modification of beta-casein (betaCN) by glucose and methylglyoxal (MGO). betaCN (1.33 M, 3.2%) was heated with either glucose (1.345 M, 4.6%) or MGO (1 mM) at 95 degrees C for up to 4 h. Tryptic digests were prepared and analysed by ultra performance liquid chromatography electrospray ionisation mass spectrometry (UPLC-ES/MS). The sites of formation of the Amadori product, N(epsilon)-(fructosyl)lysine (FL), and the advanced glycation end-products, N(epsilon)-(carboxymethyl)lysine (CML), MGO-derived dihydroxyimidazolidine (MG-DH) and MGO-derived hydroimidazolone (MG-HI), were located. FL and CML were detected at K107 and K176 residues in betaCN/glucose incubations. Indigenous N ( epsilon )-(lactulosyl)lysine was detected at K107 only. MG-DH and MG-HI were detected at R202 and possibly R183 residues in both betaCN/glucose and betaCN/MGO incubations. Glycation of betaCN by glucose and MGO resulted in similar site specificity for MG-DH and MG-HI formation.
在体外生理条件下，羰基化合物对蛋白质的修饰具有位点特异性。关于在与食品加工相关的加热条件下蛋白质糖基化的位点特异性的报道较少。本研究的目的是确定葡萄糖和甲基乙二醛（MGO）对β-酪蛋白（βCN）修饰的位点特异性。将βCN（1.33 M，3.2%）与葡萄糖（1.345 M，4.6%）或MGO（1 mM）在95℃下加热长达4小时。制备胰蛋白酶消化物并通过超高效液相色谱电喷雾电离质谱（UPLC-ES/MS）进行分析。确定了阿马多里产物N（ε）-（果糖基）赖氨酸（FL）以及晚期糖基化终产物N（ε）-（羧甲基）赖氨酸（CML）、MGO衍生的二羟基咪唑烷（MG-DH）和MGO衍生的氢咪唑酮（MG-HI）的形成位点。在βCN/葡萄糖孵育物中的K107和K176残基处检测到FL和CML。仅在K107处检测到内源性N（ε）-（乳果糖基）赖氨酸。在βCN/葡萄糖和βCN/MGO孵育物的R202以及可能的R183残基处检测到MG-DH和MG-HI。葡萄糖和MGO对βCN的糖基化导致MG-DH和MG-HI形成具有相似的位点特异性。
