Platform development for expression and purification of stable isotope labeled monoclonal antibodies in Escherichia coli
The widespread use of monoclonal antibodies (mAbs) as a platform for therapeutic drug development in the pharmaceutical industry has led to an increased interest in robust experimental approaches for assessment of mAb structure, stability and dynamics. The ability to enrich proteins with stable isot...
Saved in:
Published in | mAbs Vol. 10; no. 7; pp. 992 - 11 |
---|---|
Main Authors | , , , , , , , |
Format | Journal Article |
Language | English |
Published |
United States
Taylor & Francis
01.10.2018
|
Subjects | |
Online Access | Get full text |
Cover
Loading…
Summary: | The widespread use of monoclonal antibodies (mAbs) as a platform for therapeutic drug development in the pharmaceutical industry has led to an increased interest in robust experimental approaches for assessment of mAb structure, stability and dynamics. The ability to enrich proteins with stable isotopes is a prerequisite for the in-depth application of many structural and biophysical methods, including nuclear magnetic resonance (NMR), small angle neutron scattering, neutron reflectometry, and quantitative mass spectrometry. While mAbs can typically be produced with very high yields using mammalian cell expression, stable isotope labeling using cell culture is expensive and often impractical. The most common and cost-efficient approach to label proteins is to express proteins in Escherichia coli grown in minimal media; however, such methods for mAbs have not been reported to date. Here we present, for the first time, the expression and purification of a stable isotope labeled mAb from a genetically engineered E. coli strain capable of forming disulfide bonds in its cytoplasm. It is shown using two-dimensional NMR spectral fingerprinting that the unlabeled mAb and the mAb singly or triply labeled with
C,
N,
H are well folded, with only minor structural differences relative to the mammalian cell-produced mAb that are attributed to the lack of glycosylation in the Fc domain. This advancement of an E. coli-based mAb expression platform will facilitate the production of mAbs for in-depth structural characterization, including the high resolution investigation of mechanisms of action. |
---|---|
Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Color versions of one or more of the figures in the article can be found online at www.tandfonline.com/kmab. |
ISSN: | 1942-0862 1942-0870 |
DOI: | 10.1080/19420862.2018.1496879 |