Stefan Stahl Per Ake Nygren and Mathias Uhlen 1 Introduction

The strong and specific interaction between the Fc part of IgG and staphylococcal protein A (SpA) was utilized by Uhlen and coworkers (I) to create the first described system allowing affinity purification of expressed gene products. To date, a multitude of proteins have been produced as fusions to the IgG-bindmg domains of staphylococcal protein A, in several different hosts such as gram-positive and gram-negative bacteria (2,3), yeast (4), CHO cells (5), insect cells (6) and plants (7).

SpA is an immunoglobulin-binding surface receptor found on the gram-positive bacterium Staphylococcus aureus The SpA, which has found extensive use in immunological and biotechnological research (8—13), binds to the constant (Fc) part of certain immunoglobulins, but the exact biological significance of this property is not clear (14). Functional and structural analysis of SpA has revealed the presence of a signal peptide, S, that is processed during secretion; five highly homologous domains: E, D, A, B, and C, capable of binding to IgG (15); and a cell wall attaching structure, designated XM (16-18) (Fig. 1). Here, X represents a charged and repetitive region, postulated to interact with the peptidoglycan cell wall (16), whereas M is a region common for gram-positive cell surface bound receptors containing an LPXaaTGXaa motif, linked to a C-terminal hydrophobic region ending with a charged tail (17,18). It has been demonstrated that all three regions are required for cell surface anchoring (17) and it has been suggested that the cell wall sorting is accompanied by proteolytic cleavage at the SpA C-terminus and covalent linking of the surface receptor to the cell wall (18). SpA binds to IgG from most mammalian species, including man. Of the four subclasses of human IgG, SpA binds to IgGl, IgG2, and IgG4 but shows only weak interaction with IgG3 (19).

From Methods in Molecular Biology, vol 63 Recombinant Protein Protocols Detection and Isolation Edited by R Tuan Humana Press Inc , Totowa, NJ

IgG-binding domains








57 kDa

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