Buffered non-fermenter system for lab-scale production of secreted recombinant His-tagged proteins in Saccharomyces cerevisiae. Biotechniques 2002 Dec;33(6):1296-300
Date
12/31/2002Pubmed ID
12503315DOI
10.2144/02336pt02Scopus ID
2-s2.0-0036913195 (requires institutional sign-in at Scopus site) 10 CitationsAbstract
Expression of recombinant proteins using a secretion system can minimize co-purification of contaminating host proteins. Production of His-tagged recombinant proteins in the yeast alpha-factor secretion system has previously required a fermenter system to control the growth conditions such as pH of the yeast culture. We describe an inexpensive non-fermenter system for the production of secreted recombinant His-tagged proteins in Saccharomyces cerevisiae that uses a buffered low peptone YP glycerol medium, which does not interfere with immobilized metal affinity chromatography. Maspin, a tumor suppressor serpin, was expressed as a secreted N-terminal His/FLAG-tagged protein. Purification of the soluble active recombinant protein only requires centrifugation, concentration by ultrafiltration, and Ni2+ affinity chromatography. Purified protein yields of this system are 3-5 mg/L culture medium.
Author List
Ngamkitidechakul C, Twining SSAuthor
Sally S. Twining PhD Assistant Dean, Professor in the Biochemistry department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
BuffersCentrifugation
Chromatography, Affinity
Cloning, Molecular
Culture Media
Genes, Tumor Suppressor
Histidine
Humans
Hydrogen-Ion Concentration
Nickel
Nitrilotriacetic Acid
Oligopeptides
Peptides
Protein Biosynthesis
Proteins
Recombinant Fusion Proteins
Saccharomyces cerevisiae
Serpins
Ultrafiltration
