Maltose binding protein (MBP) fusion proteins with low or no affinity to amylose resins can be single-step purified using a novel anti-MBP monoclonal antibody.
The maltose binding protein (MBP) fusion protein expression system is a powerful tool to produce and isolate recombinant proteins in E. coli. Whereas the conventional isolation technique for MBP-fusion proteins takes advantage of the binding affinity of MBP to maltose, this method is limited insofar as the biological activity of MBP has to be fully conserved for a successful purification. In this study, a novel monoclonal antibody (mAb) specific for MBP, termed HAM-19, was generated and its application in the purification and detection of MBP-fusion proteins determined. Using anti-MBP immunoaffinity columns, even recombinant MBP fusion products with lowered or impaired binding affinity to maltose were purified in a single step procedure. In comparison to amylose resins, HAM-19 immunoaffinity columns showed a higher binding capacity and affinity to MBP-fusion proteins. Furthermore, the mAb HAM-19 also provides a technical improvement over polyclonal antisera for the detection and analysis of MBP-fusion proteins which are under use in various forms in the fields of molecular and cellular biology.
Park
J H
JH
Peptide Engineering Research Unit, Korea Research Institute of Bioscience and Biotechnology, KIST, Taejon.
Choi
E A
EA
Cho
E W
EW
Hahm
K S
KS
Kim
K L
KL
D018528:ATP-Binding Cassette Transporters; D000688:Amylose / Q000737:chemistry; D000818:Animals; D000837:Anion Exchange Resins / Q000737:chemistry; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology*; D015153:Blotting, Western; D002352:Carrier Proteins / Q000276:immunology / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D051379:Mice; D008807:Mice, Inbred BALB C; D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D012685:Sepharose / Q000737:chemistry
D018528:ATP-Binding Cassette Transporters; D000837:Anion Exchange Resins; D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins; C461599:maltose transport system, E coli; D000688:Amylose; D012685:Sepharose