loading page

Single Step Purification of a Small Non-mAb Biologic by Peptide-ELP based Affinity Precipitation
  • +4
  • Akshat Mullerpatan,
  • Ronit Ghosh,
  • Erin Kane,
  • André Nascimento,
  • Henrik Andersen,
  • Steven Cramer,
  • Pankaj Karande
Akshat Mullerpatan
Rensselaer Polytechnic Institute
Author Profile
Ronit Ghosh
Rensselaer Polytechnic Institute
Author Profile
Erin Kane
Rensselaer Polytechnic Institute
Author Profile
André Nascimento
Rensselaer Polytechnic Institute
Author Profile
Henrik Andersen
Bristol-Myers Squibb Co
Author Profile
Steven Cramer
Rensselaer Polytechnic Institute
Author Profile
Pankaj Karande
Rensselaer Polytechnic Institute
Author Profile

Peer review status:IN REVISION

03 May 2020Submitted to Biotechnology and Bioengineering
04 May 2020Assigned to Editor
04 May 2020Submission Checks Completed
26 May 2020Reviewer(s) Assigned
20 Jun 2020Review(s) Completed, Editorial Evaluation Pending
20 Jun 2020Editorial Decision: Revise Minor

Abstract

Affinity precipitation using stimulus-responsive biopolymers such as Elastin-like Polypeptides (ELPs) have been successfully employed for the purification of monoclonal antibodies. In the current work, we extend these studies to the development of an ELP-peptide fusion for the affinity precipitation of the therapeutically relevant small non-mAb biologic, AdP. A 12-mer affinity peptide ligand (P10) was identified by a primary phage biopanning followed by a secondary in-solution fluorescence polarization screen. Peptide P10 and AdP interacted with a KD of 19.5 µM. A fusion of P10 with ELP was then shown to be successful in selectively capturing the biologic from a crude mixture. While pH shifts alone were not sufficient for product elution, the use of pH in concert with fluid phase modifiers such as NaCl, arginine or ethylene glycol was successful. In particular, the use of pH 8.5 and an arginine concentration of 500 mM enabled > 80% product recovery. The overall process performance evaluated by SDS-PAGE and reversed-phase UPLC analyses, indicated the successful single-step purification of the biologic from an E. coli lysate resulting in ~90% purity and >80% recovery. These results demonstrate that phage display can be readily employed to identify a peptide ligand capable of successfully carrying out the purification of a non-antibody biological product using ELP-based affinity precipitation.