Shortened engineered human antibody CH2 domains increased stability and binding to the human neonatal Fc receptor

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Status	已發表Published
	Shortened engineered human antibody CH2 domains increased stability and binding to the human neonatal Fc receptor
	Gong, R.; Wang, Y.; Feng, Y.; Zhao, Q.; Dimitrov, D.
	2011-05-05
Source Publication	Journal of Biological Chemistry
ISSN	0021-9258
Pages	27288-27293
Abstract	The immunoglobulin (Ig) constant CH2 domain is critical for antibody effector functions. Isolated CH2 domains are promising scaffolds for construction of libraries containing diverse binders that could also confer some effector functions. We have shown previously that an isolated human CH2 domain is relatively unstable to thermally induced unfolding, but its stability can be improved by engineering an additional disulfide bond (Gong, R., Vu, B. K., Feng, Y., Prieto, D. A., Dyba, M. A., Walsh, J. D., Prabakaran, P., Veenstra, T. D., Tarasov, S. G., Ishima, R., and Dimitrov, D. S. (2009) J. Biol. Chem. 284, 14203-14210). We have hypothesized that the stability of this engineered antibody domain could be further increased by removing unstructured residues. To test our hypothesis, we removed the seven N-terminal residues that are in a random coil as suggested by our analysis of the isolated CH2 crystal structure and NMR data. The resulting shortened engineered CH2 (m01s) was highly soluble, monomeric, and remarkably stable, with a melting temperature (T(m)) of 82.6 °C, which is about 10 and 30 °C higher than those of the original stabilized CH2 (m01) and CH2, respectively. m01s and m01 were more resistant to protease digestion than CH2. A newly identified anti-CH2 antibody that recognizes a conformational epitope bound to m01s significantly better (>10-fold higher affinity) than to CH2 and slightly better than to m01. m01s bound to a recombinant soluble human neonatal Fc receptor at pH 6.0 more strongly than CH2. These data suggest that shortening the m01 N terminus significantly increases stability without disrupting its conformation and that our approach for increasing stability and decreasing size by removing unstructured regions may also apply to other proteins.
Keyword	Antibodies CH2 engineered antibody domains Fc neonatal receptor
URL	View the original
Language	英語English
The Source to Article	PB_Publication
PUB ID	55342
Document Type	Journal article
Collection	DEPARTMENT OF BIOMEDICAL SCIENCES
Corresponding Author	Dimitrov, D.
Recommended Citation GB/T 7714	Gong, R.,Wang, Y.,Feng, Y.,et al. Shortened engineered human antibody CH2 domains increased stability and binding to the human neonatal Fc receptor[J]. Journal of Biological Chemistry, 2011, 27288-27293.
APA	Gong, R.., Wang, Y.., Feng, Y.., Zhao, Q.., & Dimitrov, D. (2011). Shortened engineered human antibody CH2 domains increased stability and binding to the human neonatal Fc receptor. Journal of Biological Chemistry, 27288-27293.
MLA	Gong, R.,et al."Shortened engineered human antibody CH2 domains increased stability and binding to the human neonatal Fc receptor".Journal of Biological Chemistry (2011):27288-27293.

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