Catalytic digestion of human tumor necrosis factor-α by antibody heavy chain

It has long been an important task to prepare a catalytic antibody capable of digesting a targeting crucial protein that controls specific life functions. Tumor necrosis factor-alpha (TNF-alpha) is a cytokine and an important molecule concerned with autoimmune diseases such as rheumatoid arthritis, chronic obstructive pulmonary disease, and Crohn's disease. A mAb (ETNF-6 mAb) raised against human TNF-alpha was prepared, and the steric conformation was created by using molecular modeling after the cDNA was sequenced. The heavy chain (ETNF-6-H) of the mAb was considered to possess a catalytic triad-like structure in the complementarity determining regions (CDRs). As a result, ETNF-6-H exhibited a peptidase and a protease activity. In fact, ETNF-6-H predominantly cleaved the Ser5-Arg6 bond of TNF-alpha at the first step, resulting in the generation of a fragment of similar to 17 kDa. This fragment was digested to a smaller molecule of 15 kDa by scission of the Gln21-Ala22 bond. The intermediate product was further converted into a fragment of 13.3 kDa by successive cleavage of the Leu36-Leu37 and Asn39-Gly40 bonds. The heavy chain possessed a protease activity against TNF-alpha with a multicleavage site.