ALPHA,OMEGA-BIS-QUATERNARY AMMONIUM ALKANES AS EFFECTIVE BUFFER ADDITIVES FOR ENHANCED CAPILLARY ELECTROPHORETIC SEPARATION OF GLYCOPROTEINS

被引:48
作者
ODA, RP [1 ]
MADDEN, BJ [1 ]
SPELSBERG, TC [1 ]
LANDERS, JP [1 ]
机构
[1] MAYO CLIN & MAYO FDN,DEPT BIOCHEM & MOLEC BIOL,ROCHESTER,MN 55905
基金
英国医学研究理事会;
关键词
D O I
10.1016/0021-9673(94)80055-3
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The egg white glycoprotein, ovalbumin, is known to be microheterogeneous as a result of its varied glycan content. The use of 1,4-diaminobutane (DAB) as a buffer additive has been shown to be key in the high-resolution capillary electrophoretic separation of ''glycoforms'' of this protein [Anal. Biochem. 205 (1992) 115]. Although a separation buffer consisting of 100 mM berate and 1 mM DAB allowed for adequate separation of ovalbumin glycoforms, prolonged separation times of 35-45 min were undesirable. In the present study, the alpha,omega-bisquaternary ammonium alkanes, hexamethonium bromide (C(6)MetBr), hexamethonium chloride (C(6)MetC1) and decamethonium bromide (C(10)MetBr) were tested as buffer additives for their effectiveness in the separation of ovalbumin glycoforms. Where 1 mM DAB gave optimal separation in ca. 45 min, 100 mu M C(6)MetCl or C(10)MetBr yielded comparable resolution in less than 20 min. Results with the C(10)MetBr were better than those obtained with C(6)MetBr, indicating that there may be a correlation between effectiveness and alkyl chain length. Use of the chloride salt of C(6)Met afforded the same resolution as the bromide salt in slightly shorter analysis time. The rank order for their effectiveness was found to be C(10)MetBr > C(6)MetCl > C(6)MetBr > DAB. These results allow for speculation on the mode through which these additives exert their effect on resolution. Included in these are additive-wall coating interactions, protein-additive interactions, protein-wall interactions or any combination of these.
引用
收藏
页码:85 / 92
页数:8
相关论文
共 27 条
[21]  
TAI T, 1975, J BIOL CHEM, V250, P8569
[22]   CAPILLARY ELECTROPHORETIC SEPARATIONS OF PROTEINS USING NONIONIC SURFACTANT COATINGS [J].
TOWNS, JK ;
REGNIER, FE .
ANALYTICAL CHEMISTRY, 1991, 63 (11) :1126-1132
[23]   SEPARATION OF CARBOHYDRATE-MEDIATED MICROHETEROGENEITY OF RECOMBINANT-HUMAN-ERYTHROPOIETIN BY FREE SOLUTION CAPILLARY ELECTROPHORESIS - EFFECTS OF PH, BUFFER TYPE AND ORGANIC ADDITIVES [J].
TRAN, AD ;
PARK, SG ;
LISI, PJ ;
HUYNH, OT ;
RYALL, RR ;
LANE, PA .
JOURNAL OF CHROMATOGRAPHY, 1991, 542 (02) :459-471
[24]   CAPILLARY ELECTROPHORETIC SEPARATION OF HUMAN RECOMBINANT ERYTHROPOIETIN (RHUEPO) GLYCOFORMS [J].
WATSON, E ;
YAO, F .
ANALYTICAL BIOCHEMISTRY, 1993, 210 (02) :389-393
[25]   SEPARATION OF CATIONIC PROTEINS VIA CHARGE REVERSAL IN CAPILLARY ELECTROPHORESIS [J].
WIKTOROWICZ, JE ;
COLBURN, JC .
ELECTROPHORESIS, 1990, 11 (09) :769-773
[26]   USE OF HIGH-PERFORMANCE CAPILLARY ELECTROPHORESIS TO MONITOR CHARGE HETEROGENEITY IN RECOMBINANT-DNA DERIVED PROTEINS [J].
WU, SL ;
TESHIMA, G ;
CACIA, J ;
HANCOCK, WS .
JOURNAL OF CHROMATOGRAPHY, 1990, 516 (01) :115-122
[27]  
YAMASHITA K, 1978, J BIOL CHEM, V253, P3862