Solution properties of Escherichia coli-expressed VH domain of anti-neuraminidase antibody NC41

Alexander A. Kortt, Robin E. Guthrie, Mark G. Hinds, Barbara E. Power, Neva Ivancic, J. Bruce Caldwell, L. Clem Gruen, Raymond S. Norton, Peter J. Hudson

Research output: Contribution to journalArticleResearchpeer-review

26 Citations (Scopus)


The VH domain of anti-influenza neuraminidase antibody NC41, with and without a C-terminal hydrophilic marker peptide (FLAGTM), has been expressed in high yield (15-27 mg/L) in Escherichia coli. Both forms were secreted into the periplasm where they formed insoluble aggregates which were solubilized quantitatively with 2 M guanidine hydrochloride and purified to homogeneity by ion-exchange chromatography. The VH-FLAG was composed of three isoforms (pI values of ∼4.6, 4.9, and 5.3) and the VH molecule was composed of two isoforms with pI values of 5.1 and 6.7; the difference between the VH isoforms was shown to be due to cyclization of the N-terminal glutamine residue in the pI 5.1 isoform. At 20°C and concentrations of 5-10mg/ml the VH domain dimerized in solution and then partly precipitated, resulting in the broadening of resonances in its1H NMR spectrum. Reagents such as CHAPS, n-octylglucoside, and ethylene glycol, which presumably mask the exposed hydrophobic interface of the VH molecule, prevented dimerization of the VH and permitted good-quality NMR spectra on isotope-labeled protein to be obtained.

Original languageEnglish
Pages (from-to)167-178
Number of pages12
JournalJournal of Protein Chemistry
Issue number3
Publication statusPublished - Apr 1995
Externally publishedYes


  • Antibody
  • detergent stabilization of monomer
  • dimerization
  • NMR analysis
  • V domain

Cite this