Principal Investigator (PI): Dr. Jenny
Ottosson
Personnel: Johanna Steen (PhD-student),
Hanna Tegel (PhD-student), Anna Konrad (research engineer),
Asif Halimi (technician) and Ulla Wrethagen (technician)
Responsibility: To express and purify PrEST-proteins
to use for the preparation of antigens and affinity columns.
Description: The Protein Factory group is
responsible for protein expression and purification in the
Human Proteome Resource program. Proteins are expressed as
His6ABP fusions in E. coli shake flask cultures upon induction
with IPTG. A fully automated protein purification system has
been developed to allow for purifications of up to 60 cell
lysates at a time. One-step purification is enabled by the
hexahistidine affinity tag and metal affinity chromatography
(IMAC) and performed under denaturing conditions. After evaluation
of protein concentration, purity, and solubility the molecular
weight of the PrEST proteins are determined by electrospray
mass spectrometry as a final quality control. The purified
proteins are then used to prepare antigens and affinity columns
with PrEST-ligands. In addition Protein Factory produces affinity
resin with His6ABP-ligand.
Abbreviations:
ABP - Albumin Binding Protein
IPTG - Isopropyl-B-D-Thiogalactopyranoside
IMAC - Immobilized Metal Affinity Chromatography
