title: Easy and efficient protocol for purification of recombinant peptides.
abstract: Peptide synthesis and purification remains a challenge. Low abundance leads to small yields when peptides are purified from natural sources. On the other hand, synthetic methods are limited by the chemical properties of the amino acids and the concurrent aggregation of peptides. In this paper, we report a versatile, high yielding and general purification method for randomly chosen recombinant peptides of variable sizes (ranging from ∼1.7kDa to ∼10kDa). Expressed as fusion proteins with commonly used tag proteins, these peptides are cleaved by 'PreScission protease' in a volatile buffer that makes concentration and recovery of the peptide easy. Separation of the cleaved peptide is achieved by selective precipitation of the larger tag protein with acetonitrile; leaving the peptide in solution. Our protocol can be used to generate a wide variety of peptides in significant quantities for biochemical, biophysical and physiological studies.
authors.affiliation: Department of Biotechnology, IIT Madras, Chennai 600 036, India. Electronic address: candidprashant@gmail.com.
authors.forename: Prashant
authors.initials: P
authors.lastname: Kumar
authors.affiliation: Department of Biotechnology, IIT Madras, Chennai 600 036, India. Electronic address: agk@iitm.ac.in.
authors.forename: Gopala K
authors.initials: GK
authors.lastname: Aradhyam
mesh_terms: D000097:Acetonitriles; D000818:Animals; D001709:Biotechnology; D004926:Escherichia coli; D005982:Glutathione Transferase; D006801:Humans; D010447:Peptide Hydrolases; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D000097:Acetonitriles; D010455:Peptides; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D010447:Peptide Hydrolases; C032159:acetonitrile
keywords: Acetonitrile; Glutathione S-transferase (GST); Peptides; PreScission protease; Volatile buffer