Comparison of three distinct ELLA protocols for determination of apparent affinity constants between Con A and glycoproteins

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• 作者：D. Mislovič ; ová ; ^a ; J. Katrlí ; k^a ; E. Paulovič ; ová ; ^b ; P. Gemeiner^a ; J. Tkac^a ; ^{Jan.Tkac@savba.sk}
• 关键词：Concanavalin A ; Glycoproteins ; ELLA ; Lectin ; Apparent affinity constants
• 刊名：Colloids and Surfaces B ; Biointerfaces
• 出版年：2012
• 期刊代码：14_09277765
• 类别：ms
• 出版时间：1 June, 2012
• 卷：94
• 期：Complete
• 页码：163-169
• 文件大小：518 K

摘要

A procedure for determination of apparent affinity constants between Concanavalin A (Con A) and naturally d-mannose containing glycoproteins using enzyme-linked lectin assay (ELLA) is reported. Three distinct ELLA protocols are compared to each other with 3 different fitting models used (Liliom, Hill with and without a cooperativity factor). The glycoproteins were physisorbed on a highly charged polystyrene solid surface of immunoassay plates and the amount of lectin bound to the glycoproteins was determined by photometry. The interactions of Con A with five mannose-containing glycoproteins, invertase (INV), glucoamylase (GA), glucose oxidase (GOx), ovalbumin (OVA), and transferrin (TRF) were quantified with apparent affinity constant being in the range 2 脳 10^鈭? to 9 脳 10^鈭? M. The strength of interaction between Con A and glycoproteins is discussed on the basis of glycan structure/exposure on the protein backbone for each glycoprotein.