title: Isomerization of Asp-Asp motif in model peptides and a monoclonal antibody Fab fragment.
abstract: Isomerization of aspartyl (Asp or D) residues is a critical degradation route to consider for stable monoclonal antibody formulations. Among the known hotspot sequences, the DD motif is relatively understudied. To gain mechanistic insights, we used model hexapeptides, YADXFK, YADDXK, and DIDDDM, as surrogates for the hotspots in a Fab protein (YADDFK and DIDDDM), to characterize the rate-pH profile of Asp isomerization. Compared with the YADGFK peptide, isomerization of D3 (the first D in the DD pair) in YADDFK was highly pH dependent. Comparison of rate-pH profiles of YADDFK, YADNFK, and YADHFK revealed a charge effect of the n + 1 residue-isomerization rate is accelerated by the positive side chain and reduced by negative side chain at n + 1 residue. Studies on YADDFK, YADDAK, and YADDGK indicated a mutual impact of D3 and D4 on their respective isomerization rates through charge effect. Comparison of rate-pH profile of DIDDDM sequence in peptide models with that in the complementary determining region of the Fab showed a faster rate in the Fab than in peptides, presumably because of contribution from structural factors in the former.
authors.affiliation: Early Stage Pharmaceutical Development, Genentech, South San Francisco, California 94080, USA. yi.li@gene.com
authors.forename: Li
authors.initials: L
authors.lastname: Yi
authors.forename: Nia
authors.initials: N
authors.lastname: Beckley
authors.forename: Benson
authors.initials: B
authors.lastname: Gikanga
authors.forename: Jennifer
authors.initials: J
authors.lastname: Zhang
authors.forename: Y John
authors.initials: YJ
authors.lastname: Wang
authors.forename: Hung-Wei
authors.initials: HW
authors.lastname: Chih
authors.forename: Vikas K
authors.initials: VK
authors.lastname: Sharma
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D061067:Antibodies, Monoclonal, Humanized / Q000737:chemistry; D004151:Dipeptides / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D007536:Isomerism; D007700:Kinetics
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D004151:Dipeptides; D007140:Immunoglobulin Fab Fragments; C108952:aspartyl-aspartic acid