Improving solubility and refolding efficiency of human V(H)s by a novel mutational approach

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DOIResolve DOI: http://doi.org/10.1093/protein/gzl037
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TypeArticle
Journal titleProtein Engineering, Design & Selection
Volume19
Issue11
Pages503509; # of pages: 7
SubjectHuman; Mutation; pha; Solubility; Temperature
AbstractThe antibody V(H) domains of camelids tend to be soluble and to resist aggregation, in contrast to human V(H) domains. For immunotherapy, attempts have therefore been made to improve the properties of human V(H)s by camelization of a small set of framework residues. Here, we have identified through sequence comparison of well-folded llama V(H) domains an alternative set of residues (not typically camelid) for mutation. Thus, the solubility and thermal refolding efficiency of a typical human V(H), derived from the human antibody BT32/A6, were improved by introduction of two mutations in framework region (FR) 1 and 4 to generate BT32/A6.L1. Three more mutations in FR3 of BT32/A6.L1 further improved the thermal refolding efficiency while retaining solubility and cooperative melting profiles. To demonstrate practical utility, BT32/A6.L1 was used to construct a phage display library from which were isolated human V(H)s with good antigen binding activity and solubility. The engineered human V(H) domains described here may be useful for immunotherapy, due to their expected low immunogenicity, and in applications involving transient high temperatures, due to their efficient refolding after thermal denaturation
Publication date
LanguageEnglish
AffiliationNRC Biotechnology Research Institute; National Research Council Canada; NRC Institute for Biological Sciences
Peer reviewedNo
NRC number42510
NPARC number3539224
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Record identifierd2649e55-49a8-457e-9aa8-9a8563246ebb
Record created2009-03-01
Record modified2016-05-09
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