A three-step purification strategy for isolation of hamster TIG2 from CHO cells: characterization of two processed endogenous forms.
We have recently isolated a bioactive, circulating protein of human tazarotene-induced gene-2 (TIG2) as the natural ligand of the orphan receptor ChemR23. Here we describe a simplified method for the isolation of hamster TIG2 protein from Chinese hamster ovary (CHO) cell supernatant. Using a heparin-affinity column followed by two reversed phase chromatography steps resulted in the isolation of pure biologically active material. Two processed bioactive forms of Chinese hamster TIG2 were identified by Edman sequencing and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF-MS) mass fingerprint analysis, representing the amino acid residues T20 to F156, and T20 to A155 of the 163 amino acid propeptide. Comparison with the predicted aa-sequence indicates a mutation or modification within the C-terminal end of the peptide.
Busmann
Annette
A
IPF PharmaCeuticals GmbH, Feodor-Lynen-Str. 31, D-30625 Hannover, Germany.
Walden
Michael
M
Wendland
Martin
M
Kutzleb
Christian
C
Forssmann
Wolf-Georg
WG
John
Harald
H
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D016147:Genes, Tumor Suppressor; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D051381:Rats; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization