Oda R P, Prasad R, Stout R L, Coffin D, Patton W P, Kraft D L, O'Brien J F, Landers J P
Department of Laboratory Medicine and Pathology, Mayo Clinic/Foundation, Rochester, MN, USA.
Electrophoresis. 1997 Sep;18(10):1819-26. doi: 10.1002/elps.1150181017.
The heterogeneity associated with protein glycoforms has been a challenge to analytical chemists and the subject of structure-function studies for biochemists since their presence in biological systems had been confirmed some three decades ago. Initial investigations led to discoveries of synthetic and degradative pathways, and brief forays into functional determination of the "glyco" portion on the protein activity in glycoproteins. Only recently has it come to our understanding that variations from the "normal" glycosylation patterns might be indicative of pathological states. The presence of certain transferrin (Tf) glycoforms in human serum has been shown to correlate with certain clinical syndromes. Hence, the ability to separate and quantitatively measure the various forms of human Tf has become increasingly important. It this study, we demonstrate that a simple method utilizing a DB-17-coated capillary to slow endoosmotic flow and a sieving buffer containing hydroxyethyl cellulose allows for the resolution of sialoforms of transferrin. An analysis time of less than eight minutes allows for baseline resolution of the lower sialoforms of Tf, presenting a simple, rapid test for carbohydrate-deficient transferrin (CDT). We demonstrate the utility of this methodology for the facile diagnosis of carbohydrate-deficient glycoprotein syndrome, and postulate that it may allow for the detection of other carbohydrate-deficient protein-related disease states.
自从大约三十年前蛋白质糖型在生物系统中的存在得到证实以来，与蛋白质糖型相关的异质性一直是分析化学家面临的挑战，也是生物化学家进行结构-功能研究的主题。最初的研究导致了合成和降解途径的发现，并对糖蛋白中蛋白质活性上“糖”部分的功能测定进行了简要探索。直到最近我们才认识到，与“正常”糖基化模式的差异可能表明病理状态。已证明人血清中某些转铁蛋白（Tf）糖型的存在与某些临床综合征相关。因此，分离和定量测量人Tf各种形式的能力变得越来越重要。在本研究中，我们证明了一种简单的方法，即利用涂有DB-17的毛细管减缓内渗流，并使用含有羟乙基纤维素的筛分缓冲液，可以分离转铁蛋白的唾液酸化形式。不到八分钟的分析时间可以实现Tf较低唾液酸化形式的基线分离，为诊断缺糖转铁蛋白（CDT）提供了一种简单、快速的检测方法。我们证明了这种方法在糖蛋白缺陷综合征简易诊断中的实用性，并推测它可能有助于检测其他与缺糖蛋白相关的疾病状态。
