Steric Accessibility of the Cleavage Sites Dictates the Proteolytic Vulnerability of the Anti‐HIV‐1 Antibodies 2F5, 2G12, and PG9 in Plants. Issue 3 (8th November 2019)

• Record Type: Journal Article
• Authors: Puchol Tarazona, Alejandro A.; Lobner, Elisabeth; Taubenschmid, Yvonne; Paireder, Melanie; Torres Acosta, Juan A.; Göritzer, Kathrin; Steinkellner, Herta; Mach, Lukas
• Is Part Of: Biotechnology journal. Volume 15:Issue 3(2020)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2019-11-08
• Subjects: antibody degradation -- HIV -- Nicotiana benthamiana -- recombinant protein expression -- serine protease; Biotechnology -- Periodicals; 660.605
• DOI: 10.1002/biot.201900308
• Languages: English
• ISSNs: 1860-6768

Abstract

Abstract: Broadly neutralizing antibodies (bNAbs) to human immunodeficiency virus type 1 (HIV‐1) hold great promise for immunoprophylaxis and the suppression of viremia in HIV‐positive individuals. Several studies have demonstrated that plants as Nicotiana benthamiana are suitable hosts for the generation of protective anti‐HIV‐1 antibodies. However, the production of the anti‐HIV‐1 bNAbs 2F5 and PG9 in N. benthamiana is associated with their processing by apoplastic proteases in the complementarity‐determining‐region (CDR) H3 loops of the heavy chains. Here, it is shown that apoplastic proteases can also cleave the CDR H3 loop of the bNAb 2G12 when the unusual domain exchange between its heavy chains is prevented by the replacement of Ile 19 with Arg. It is demonstrated that CDR H3 proteolysis leads to a strong reduction of the antigen‐binding potencies of 2F5, PG9, and 2G12‐I19R. Inhibitor profiling experiments indicate that different subtilisin‐like serine proteases account for bNAb fragmentation in the apoplast. Differential scanning calorimetry experiments corroborate that the antigen‐binding domains of wild‐type 2G12 and 4E10 are more compact than those of proteolysis‐sensitive antibodies, thus shielding their CDR H3 regions from proteolytic attack. This suggests that the extent of proteolytic inactivation of bNAbs in plants is primarily dictated by the steric accessibility of their CDR H3 loops. Abstract : Broadly neutralizing antibodies (bNAbs) hold promise for the treatment of HIV‐positive individuals. bNAb production in plant‐based expression platforms is frequently accompanied by their proteolytic inactivation. In Nicotiana benthamiana, bNAb proteolysis appears to be mediated by endogenous subtilisin‐like serine proteases.