Purification of recombinant HBc antigen expressed in Escherichia coli and Pichia pastoris: comparison of size-exclusion chromatography and ultracentrifugation
D. Rolland et al., Purification of recombinant HBc antigen expressed in Escherichia coli and Pichia pastoris: comparison of size-exclusion chromatography and ultracentrifugation, J CHROMAT B, 753(1), 2001, pp. 51-65
Hepatitis B virus core protein (HBc) is an important serology marker of hep
atitis B infection and patient follow-up. It is an M-r 21 000 protein, whic
h has the intrinsic capacity to self-assemble as a capsid-like particle. Th
e hepatitis B core protein has been expressed in Escherichia coli and Pichi
a pastoris (three different constructions) in order to select a HBc recombi
nant antigen suitable for serodiagnosis requirements with a cost effective
downstream strategy. The expression and purification of the different forms
of recombinant HBc have been described. For the last step, ultracentrifuga
tion and size-exclusion chromatography were compared. The morphology of the
se capsids was observed using an electron microscope. Our data shows that H
Bc antigen is produced in large quantities in E. coli but some contaminants
remained which were associated with the E. coli HBc protein after ultracen
trifugation or size-exclusion chromatography; The ultracentrifugation enabl
es a higher purity of HBc antigen to be obtained than size-exclusion chroma
tography but the latter enables a higher recovery rate. P. pastoris enables
the expression and extraction of a highly purified HBc antigen suitable fo
r diagnostic purposes. (C) 2001 Elsevier Science B.V. All rights reserved.