Global assessment of protein turnover in recombinant antibody producing myeloma cells.
The global turnover rates of cellular proteins and the secretion rate of a recombinant immunoglobulin G (IgG) in a myeloma cell line, NS0, were determined using SILAC proteomic analysis. After complete labeling of cellular proteins with (13)C(6), (15)N(4)-arginine, cells were transferred to unlabeled medium and the decay of the labeled arginine in proteins was monitored during exponential cell growth. After PAGE separation and mass-spectrometric identification of proteins, those detected with high confidence over at least three time points were used for the determination of turnover rates. Among the 224 proteins quantified with a protein half-life, about 15% have a degradation rate constant lower than one-tenth of specific growth rate. For most proteins, the turnover rate is insignificant in its overall dynamics. Only 6.3% of proteins have a half-life shorter than the cell doubling time. For IgG secretion, both heavy and light chain molecules follow the same kinetic behavior with a half-life estimated to be 2h. The label decay curve appears to show a second region with very slow kinetics, raising the possibility of two populations of IgG molecules with different secretion characteristics.
Yee
Joon Chong
JC
Department of Chemical Engineering and Materials Science, University of Minnesota, Minneapolis, MN 55455-0132, USA.
Jacob
Nitya M
NM
Jayapal
Karthik P
KP
Kok
Yee-Jiun
YJ
Philp
Robin
R
Griffin
Timothy J
TJ
Hu
Wei-Shou
WS
D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D002460:Cell Line; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000378:metabolism*; D007553:Isotope Labeling / Q000379:methods; D013058:Mass Spectrometry / Q000379:methods*; D008657:Metabolic Clearance Rate; D009101:Multiple Myeloma / Q000378:metabolism*; D009194:Myeloma Proteins / Q000378:metabolism*; D010449:Peptide Mapping / Q000379:methods*; D020543:Proteome / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis*
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D009194:Myeloma Proteins; D020543:Proteome; D011994:Recombinant Proteins