A novel gene fusion system to express and purify small recombinant proteins in Escherichia coli has been constructed. The concept allows for affinity purification of soluble gene products by sequential albumin- and Zn2(+)-affinity chromatography. The dual-affinity system is well suited for expression of unstable proteins as only full-length protein is obtained after purification and proteins gain proteolytic stability in the fusion protein. Here we show that the dual-affinity approach can be used for the expression of various unstable derivatives of a single IgG-binding domain based on staphylococcal protein A. Analysis of the proteolytic stabilities and the IgG-binding properties of the different mutant proteins suggest that the model for the structure of an IgG-binding domain must be re-evaluated.