Analysis of γ-carboxyglutamic acid content of protein, urine, and plasma by capillary electrophoresis and laser-induced fluorescence

被引:12
作者
Britz-Mckibbin, P
Vo, HC
MacGillivray, RTA
Chen, DDY
机构
[1] Univ British Columbia, Dept Chem, Vancouver, BC V6T 1Z1, Canada
[2] Univ British Columbia, Dept Biochem & Mol Biol, Vancouver, BC V6T 1Z3, Canada
关键词
D O I
10.1021/ac981241g
中图分类号
O65 [分析化学];
学科分类号
070302 ; 081704 ;
摘要
When the properties of an analyte are known, the separation system can be designed to make the analyte of interest migrate at either a much faster ora much slower velocity compared to other molecules in the sample matrix. A simple and sensitive method to analyze the gamma-carboxyglutamic acid (Gla) content of protein, urine, and plasma was developed using capillary electrophoresis with laser-induced fluorescence detection (CE-IIF), The separation method is designed according to the specific properties of three amino acids of interest. The number of Gla residues from three vitamin K-dependent proteins were estimated by quantifying the amount of fluorescein thiocarbamyl derivative of Gla after alkaline hydrolysis and fluorescein isothiocyanate labeling. Human prothrombin, blood coagulation factor X, and bovine osteocalcin were calculated to have 10.0 +/- 0.7, 11.0 +/- 0.6, and 2.1 +/- 0.1 Gla residues. per mole of protein, respectively, which agreed well with amino acid sequencing data. The analysis of free Gla content in urine and plasma was also demonstrated by this method. It was demonstrated that submicrograms of protein can be characterized by CE-LIF.
引用
收藏
页码:1633 / 1637
页数:5
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