A Practical Method for Uniform Isotopic Labeling of Recombinant Proteins in Mammalian Cells

Andrew P. Hansen; Andrew M. Petros; Andrew P. Mazar; Terry M. Pederson; Annemarie Rueter; Stephen W. Fesik

doi:10.1021/bi00166a001

A Practical Method for Uniform Isotopic Labeling of Recombinant Proteins in Mammalian Cells

Andrew P. Hansen, Andrew M. Petros, Andrew P. Mazar, Terry M. Pederson, Annemarie Rueter, Stephen W. Fesik

Pharmacology

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71 Scopus citations

Abstract

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.

Original language	English (US)
Pages (from-to)	12713-12718
Number of pages	6
Journal	Biochemistry
Volume	31
Issue number	51
DOIs	https://doi.org/10.1021/bi00166a001
State	Published - Feb 1 1992

ASJC Scopus subject areas

Biochemistry

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@article{0372e420ae354a5ebc041d87240a8f06,

title = "A Practical Method for Uniform Isotopic Labeling of Recombinant Proteins in Mammalian Cells",

abstract = "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.",

author = "Hansen, {Andrew P.} and Petros, {Andrew M.} and Mazar, {Andrew P.} and Pederson, {Terry M.} and Annemarie Rueter and Fesik, {Stephen W.}",

year = "1992",

month = feb,

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doi = "10.1021/bi00166a001",

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journal = "Biochemistry",

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T1 - A Practical Method for Uniform Isotopic Labeling of Recombinant Proteins in Mammalian Cells

AU - Hansen, Andrew P.

AU - Petros, Andrew M.

AU - Mazar, Andrew P.

AU - Pederson, Terry M.

AU - Rueter, Annemarie

AU - Fesik, Stephen W.

PY - 1992/2/1

Y1 - 1992/2/1

N2 - 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.

AB - 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.

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JO - Biochemistry

JF - Biochemistry

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ER -

A Practical Method for Uniform Isotopic Labeling of Recombinant Proteins in Mammalian Cells

Abstract

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