Journal
PROTEIN EXPRESSION AND PURIFICATION
Volume 134, Issue -, Pages 114-124Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.pep.2017.04.008
Keywords
Affinity purification; Antibody mimetics; nanoCLAMPs; Immunoaffinity; Resins; Native elution
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Funding
- Nectagen, Inc.
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The purification of functional proteins is a critical pre-requisite for many experimental assays. Immunoaffinity chromatography, one of the fastest and most efficient purification procedures available, is often limited by elution conditions that disrupt structure and destroy enzymatic activity. To address this limitation, we developed polyol-responsive antibody mimetics, termed nanoCLAMPs, based on a 16 kDa carbohydrate binding module domain from Clostridium perfringens hyaluronidase. nanoCLAMPs bind targets with nanomolar affinity and high selectivity yet release their targets when exposed to a neutral polyol-containing buffer, a composition others have shown to preserve quaternary structure and enzymatic activity. We screened a phage display library for nanoCLAMPs recognizing several target proteins, produced affinity resins with the resulting nanoCLAMPs, and successfully purified functional target proteins by single-step affinity chromatography and polyol elution. To our knowledge, nanoCLAMPs constitute the first antibody mimetics demonstrated to be polyol-responsive. (C) 2017 The Authors. Published by Elsevier Inc.
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