EXPRESSION AND SECRETION OF FUNCTIONAL MINIANTIBODIES MCPC603SCFVDHLXIN CELL-WALL-LESS L-FORM STRAINS OF PROTEUS-MIRABILIS AND ESCHERICHIA-COLI - A COMPARISON OF THE SYNTHESIS CAPACITIES OF L-FORM STRAINS WITH AN ESCHERICHIA-COLI PRODUCER STRAIN

The paper describes the synthesis of the phosphorylcholine-binding min iantibody McPC603scFvDhl x in cell-wall-less L-form strains of Escheri chia coli and Proteus mirabilis. Cells of these strains were transform ed with the plasmid pACK02scKan, carrying the miniantibody (miniAb) co ding sequence under the control of the lac promoter. L-form transforma nts of both species were able to synthesize the functional miniAb as a n extracellular soluble product. The highest quantities were obtained by P. mirabilis L-form strains after induction with 5 mM isopropyl bet a-D-thiogalactopyranoside (IPTG). Yields of 45-75 mg/l total antibody protein and of 10-18 mg/l functional miniAb were estimated in the grow th medium of shaking cultures 40-80 h after induction with IPTG. About 10% of the active miniAb remained cell-bound. The yields of functiona l miniAb could be optimized by lowering the growth temperature from 37 degrees C to 26-32 degrees C and by supplementation of the medium wit h 80 mM sodium fumarate. A comparison of the specific activities revea led that the P. mirabilis L-form strains have a similar synthesis capa city (2-4 mg functional miniAb/g cell dry weight) to that of the produ cer strain E. coli RV308. The results show that the processes of corre ct folding and assembling of the miniAb molecules are possible without the periplasmic compartment.