Transforming growth factor beta 1: NMR signal assignments of the recombinant protein expressed and isotopically enriched using Chinese hamster ovary cells.
The transforming growth factor beta s are a homologous family of multifunctional cytokines that regulate cell growth and differentiation. As a prelude to studies of the solution structure and dynamics of TGF-beta 1, we report virtually complete assignment of 1H and 15N resonances for this 25-kDa homodimeric protein. Recombinant TGF-beta 1 was expressed in Chinese hamster ovary cells. The cells were grown either in a completely 15N-enriched medium or in a medium containing selectively 13C, 15N-labeled amino acids to obtain either uniformly or specifically labeled protein, respectively. Two- and three-dimensional heteronuclear edited magnetic resonance spectra of the uniformly 15N-labeled protein and three samples selectively labeled with 13C and 15N yielded assignments for 96% of the backbone amide and C alpha protons and 87% of the side chain protons. To our knowledge, this is the first report of the use of an animal cell expression system to obtain extensive isotopic enrichment in order to sequentially assign a protein. The methodology described herein for the isotopic enrichment and resonance assignments of TGF-beta 1 should be generally applicable to other eukaryotic proteins expressed by animal cells.
Archer
S J
SJ
Bone Research Branch, National Institute of Dental Research, National Institutes of Health, Bethesda, Maryland 20892.
Bax
A
A
Roberts
A B
AB
Sporn
M B
MB
Ogawa
Y
Y
Piez
K A
KA
Weatherbee
J A
JA
Tsang
M L
ML
Lucas
R
R
Zheng
B L
BL
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D016466:CHO Cells; D002417:Cattle; D003001:Cloning, Molecular; D006224:Cricetinae; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry; D016212:Transforming Growth Factor beta / Q000737:chemistry* / Q000235:genetics
D000596:Amino Acids; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta