A practical method for uniform isotopic labeling of recombinant proteins in mammalian cells.

Hansen AP, Petros AM, Mazar AP, Pederson TM, Rueter A, Fesik SW
Biochemistry. 1992 31 (51): 12713-8

PMID: 1463742 · DOI:10.1021/bi00166a001

A method to obtain uniformly isotopically labeled (15N and 15N/13C) protein from mammalian cells is described. The method involves preparation of isotopically labeled media consisting of amino acids isolated from bacterial and algal extracts supplemented with cysteine and enzymatically synthesized glutamine. The approach is demonstrated by producing 15N-labeled and 15N/13C-labeled urokinase from Sp2/0 cells and successfully growing Chinese hamster ovary (CHO) cells on the labeled media. Thus, using the procedures described, isotopically labeled proteins that have been expressed in mammalian cells can be prepared, allowing them to be studied by heteronuclear multidimensional NMR techniques.

MeSH Terms (14)

Amino Acids Animals Carbon Isotopes Cells, Cultured CHO Cells Cricetinae Culture Media Cysteine Glutamine Isotope Labeling Magnetic Resonance Spectroscopy Nitrogen Isotopes Recombinant Proteins Urokinase-Type Plasminogen Activator

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