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Engineered high-affinity nanobodies recognizing staphylococcal Protein A and suitable for native isolation of protein complexes.

ABSTRACT: In addition to its high affinity for antibody Fc domains, staphylococcal Protein A has been shown to bind certain Fab domains. We investigated this in order to develop a small, recombinant Protein A-binding alternative to immunoglobulin G (IgG) from nanobodies, single-domain antibodies derived from a camelid variant IgG's variable region. We engineered a nanobody with affinity solely for Protein A as well as a dimerized version of higher affinity for typical multidomain Protein A constructs. Because this recombinant nanobody can be immobilized using a cleavable crosslinker, it has proven to be suitable for the isolation and mild elution of protein complexes in native conditions.

SUBMITTER: Fridy PC

PROVIDER: S-EPMC4404190 | biostudies-literature | 2015 May

REPOSITORIES: biostudies-literature

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Engineered high-affinity nanobodies recognizing staphylococcal Protein A and suitable for native isolation of protein complexes.

Fridy Peter C PC Thompson Mary K MK Ketaren Natalia E NE Rout Michael P MP

Analytical biochemistry 20150221

In addition to its high affinity for antibody Fc domains, staphylococcal Protein A has been shown to bind certain Fab domains. We investigated this in order to develop a small, recombinant Protein A-binding alternative to immunoglobulin G (IgG) from nanobodies, single-domain antibodies derived from a camelid variant IgG's variable region. We engineered a nanobody with affinity solely for Protein A as well as a dimerized version of higher affinity for typical multidomain Protein A constructs. Bec ...[more]

PMID: 25707320

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