Renaturation, purification and characterization of recombinant Fab-fragments produced in Escherichia coli.
Cytoplasmatic expression of murine antibody chains in Escherichia coli results in the formation of insoluble and inactive protein aggregates (inclusion bodies). By systematic variation of the parameters influencing the folding, formation of disulfide bonds and association of the constituent polypeptide chains, we have designed a renaturation procedure allowing the production of microbially expressed Fab-fragments at yields up to 40 percent of the total amount of recombinant protein. The strategy of optimization is generally applicable for disulfide containing proteins produced as inclusion bodies in bacteria. The purified recombinant antibody fragments obtained are identical with the native murine Fab in all functional and physicochemical parameters tested.
Buchner
J
J
Universität Regensburg, Institut für Biophysik and Physikalische Biochemie, FRG.
Rudolph
R
R
D000818:Animals; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D002479:Inclusion Bodies; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000096:biosynthesis*
D007140:Immunoglobulin Fab Fragments; D011994:Recombinant Proteins