Reactivity toward deamidation of asparagine residues in β‐turn structures

Abstract

Mimetics of beta-turn structures in proteins have been used to calibrate the relative reactivities toward deamidation of asparagine residues in the two central positions of a beta-turn and in a random coil. N-Acetyl-Asn-Gly-6-aminocaproic acid, an acyclic analog of a beta-turn mimic undergoes deamidation of the asparaginyl residue through a succinimide intermediate to generate N-acetyl-Asp-N-Gly-6-aminocaproic acid (6-aminocaproic acid, hereafter Aca) and N-acetyl-L-iso-aspartyl (isoAsp)-Gly-Aca (pH 8.8, 37 degrees C) approximately 3-fold faster than does the cyclic beta-turn mimic cyclo-[L-Asn-Gly-Aca] with asparagine at position 2 of the beta-turn. The latter compound, in turn, undergoes deamidation approximately 30-fold faster than its positional isomer cyclo-[Gly-Asn-Aca] with asparagine at position 3 of the beta-turn. Both cyclic peptides assume predominantly beta-turn structures in solution, as demonstrated by NMR and circular dichroism characterization. The open-chain compound and its isomer N-acetyl-Gly-Asn-Aca assume predominantly random coil structures. The latter isomer undergoes deamidation 2-fold slower than the former. Thus the order of reactivity toward deamidation is: asparagine in a random coil approximately 3x(asparagine) in position 2 of a beta-turn approximately 30x (asparagine) in position 3 of a beta-turn.

Asparagine deamidation in recombinant human lymphotoxin: Hindrance by three-dimensional structures

“…Such stabilization probably arises from structural constraints and reduced flexibility, which hinder formation of the favored conformation required for cyclic-imide formation. 24,26 Analysis of the X-ray crystal structures of the rhLT trimer and the rhLT:p55 receptor trimer-trimer complex demonstrates that the Asn 19 and Asn 40 -Asn 41 regions are both in b-turn conformations, as shown in Figures 5 and 6. Asn 19 is the third residue in a type IV or a type I b-turn, while the Asn 40 and Asn 41 residues are the second and the third residues in a type III 0 b-turn.…”

“…Fragment T3, the smallest and most hydrophilic, eluted at the solvent front, as confirmed by LC-MS. This did not allow the study of possible deamidation at the Asn 26 residue, which resides in the T3 fragment.…”

“…Studies of b-turn model peptides 26,27 and some proteins 21,28,29 have demonstrated that b-turn secondary structures can stabilize Asn against deamidation, especially when Asn is located at the third position in a b-turn structure (except for type II turns). Such stabilization probably arises from structural constraints and reduced flexibility, which hinder formation of the favored conformation required for cyclic-imide formation.…”

Mutational Analysis of a Conserved Glutamic Acid Required for Self-Catalyzed Cross-Linking of Bacteriophage HK97 Capsids

“…Deamidation of glutamine produces glutamic acid, which is the wild-type residue and so would support one cross-link for each deamidation. (17,48,52). So it seems plausible to us that the small increases above background levels in the extent of cross-linking for the Gln and Asn mutants are due to low-level side chain deamidation of these residues which can yield carboxylate-bearing side chains.…”

Synthesis, Conformational Analysis, and Biological Activity of Proposed Vitronectin Antagonists

“…Aca templates were also used to examine rates of deamidation of asparagine residues, a process of peptide degradation that produces either aspartic acid or isoaspartic acid [83]. Controlled hydrolysis of Aca-constrained dipeptides containing asparagine was carried out to correlate the position of asparagine in a ß-turn with its rate of deamidation.…”