Department of Chemistry, University of Nebraska, Lincoln, NE, USA.
Clin Chim Acta. 2011 Jan 30;412(3-4):277-85. doi: 10.1016/j.cca.2010.10.018. Epub 2010 Oct 27.
Many of the complications encountered during diabetes can be linked to the non-enzymatic glycation of proteins, including human serum albumin (HSA). However, there is little information regarding how the glycation pattern of HSA changes as the total extent of glycation is varied. The goal of this study was to identify and conduct a semi-quantitative comparison of the glycation products on HSA that are produced in the presence of various levels of glycation.
Three glycated HSA samples were prepared in vitro by incubating physiological concentrations of HSA with 15 mmol/l glucose for 2 or 5 weeks, or with 30 mmol/l glucose for 4 weeks. These samples were then digested and examined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to identify the glycation products that were formed.
It was found that the glycation pattern of HSA changed with its overall extent of total glycation. Many modifications including previously-reported primary glycation sites (e.g., K199, K281, and the N-terminus) were consistently found in the tested samples. Lysines 199 and 281, as well as arginine 428, contained the most consistently identified and abundant glycation products. Lysines 93, 276, 286, 414, 439, and 524/525, as well as the N-terminus and arginines 98, 197, and 521, were also found to be modified at various degrees of HSA glycation.
The glycation pattern of HSA was found to vary with different levels of total glycation and included modifications at the 2 major drug binding sites on this protein. This result suggests that different modified forms of HSA, both in terms of the total extent of glycation and glycation pattern, may be found at various stages of diabetes. The clinical implication of these results is that the binding of HSA to some drug may be altered at various stages of diabetes as the extent of glycation and types of modifications in this protein are varied.
糖尿病患者所遇到的许多并发症都与蛋白质的非酶糖化有关，其中包括人血清白蛋白（HSA）。然而，关于 HSA 的糖化模式如何随糖化总程度的变化而变化，目前的信息很少。本研究的目的是鉴定并对半定量比较在不同糖化水平下产生的 HSA 的糖化产物。
通过将生理浓度的 HSA 与 15mmol/L 葡萄糖孵育 2 或 5 周或与 30mmol/L 葡萄糖孵育 4 周，在体外制备了 3 个糖化 HSA 样品。然后通过基质辅助激光解吸/电离飞行时间质谱（MALDI-TOF MS）对这些样品进行消化和检查，以鉴定形成的糖化产物。
发现 HSA 的糖化模式随其总糖化程度而变化。在测试的样品中，发现了许多修饰，包括以前报道的主要糖化位点（例如 K199、K281 和 N 末端）。赖氨酸 199 和 281 以及精氨酸 428 含有最一致识别和丰富的糖化产物。赖氨酸 93、276、286、414、439 和 524/525 以及 N 末端和精氨酸 98、197 和 521 也被发现在不同程度的 HSA 糖化时被修饰。
发现 HSA 的糖化模式随总糖化程度的不同而变化，包括该蛋白质上 2 个主要药物结合位点的修饰。这一结果表明，在糖尿病的不同阶段，可能会发现不同修饰形式的 HSA，无论是在糖化总程度还是糖化模式方面。这些结果的临床意义是，随着 HSA 中糖化程度和修饰类型的变化，HSA 与某些药物的结合可能在糖尿病的不同阶段发生改变。
