BACTERIAL EXPRESSION OF A SINGLE-CHAIN FV FRAGMENT WHICH EFFICIENTLY PROTECTS THE ACETYLCHOLINE-RECEPTOR AGAINST ANTIGENIC MODULATION CAUSED BY MYASTHENIC ANTIBODIES

Monoclonal antibodies (mAb) against the main immunogenic region (MIR) of the acetylcholine receptor (AChR) are very potent in inducing antig enic modulation of the AChR in animals and in muscle cell cultures. A recombinant antibody fragment of the rat anti-MIR mAb198 was cloned by polymerase chain reaction and expressed as soluble single-chain Fv fr agment (scFv198) in E. coli and affinity purified. DNA sequencing was used to define the VH (IB) and VL (K2) chain gene usage. scFv198 was f ound immunologically and biologically active. Its binding affinity for the Torpedo AChR (K(D) = 2 +/- 0.6 nm) was very similar with that of the intact mAb198 (K(D) = 1.8 +/- 0.6 nm) while for the human AChR (K( D) = 80.7 +/- 16.6 nm) it was about four times lower than that of the intact mAb198 (K(D) = 21.6 +/- 6.6 nm). This fragment was capable of e fficiently protecting the AChR in human cell cultures, against antigen ic modulation caused by the intact mAb198 or by the antibodies from a myasthenic patient. The produced scFv198 fragment is, therefore, poten tially useful in therapeutic applications for myasthenia gravis after appropriate genetic manipulations.