Natasa Kopitar-Jerala, Franc Gubenek, Vito Turk
Recombinant Anti-Stefin A Fab Fragment: Sequence Analysis of the Variable Region and Expression in Escherichia coli
Human stefin A is an inhibitor of lysosomal cysteine
proteinases cathepsin B, H, L and S. In the present
report we describe the cloning and expression of antistefin
A Fab fragment A22 in E. coli. We have determined
the nucleotide sequences of the antibody heavy and
light chain and compared them to the murine immunoglobulin
germ line sequences. Expression of the two
antibody chains was achieved using a single vector
with a PhoA promoter and coding regions placed after
the signal sequences, directing them to the periplasmic
space. The A22 Fab fragment was extracted from
the periplasmic space and expression was confirmed
by Western blot analysis. The recombinant A22 Fab
fragment had an affinity for stefin A comparable to
the original monoclonal antibody, as determined by
ELISA.
Biological Chemistry, Walter de Gruyter
Print ISSN: 1431-6730
Volume: 381, 12/2000
Pages: 1245 - 1249
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