Amyloidogenic determinant as a substrate recognition motif of insulin‐degrading enzyme

Abstract

Insulin-degrading enzyme (IDE) is an evolutionarily conserved neutral thiol metalloprotease expressed in all mammalian tissues whose biological role is not well established. IDE has highly selective substrate specificity. It degrades insulin, glucagon, atrial natriuretic peptide, transforming growth factor K K but does not act on related hormones and growth factors. The structural properties determining whether a peptide is an IDE substrate are essentially unknown. The reported cleavage sites are not consistent with simple peptide-bond recognition and it was proposed that IDE recognizes in its substrates some elements of tertiary structure. We noticed that although IDE substrates are functionally unrelated, the majority of them share a specific property, an ability to form under certain conditions amyloid fibrils. Utilizing the residue pattern recognition procedure, this study reveals a common motif in the sequences of IDE substrates, HNHHHPSH, where H is wholly or partly hydrophobic character, N is small and neutral, P is polar, and S is polar and/or small amino acid residue. It is proposed that this sequence motif predetermines a structure recognized by IDE. The identified motif appears to be essentially the same as the proposed earlier consensus sequence for amyloid-forming peptides [Turnell and Finch, J. Mol. Biol. 227 (1992) 1205^1223]. The study suggests that IDE may play a role in elimination of potentially toxic amyloidogenic peptides.z 1998 Federation of European Biochemical Societies.

Metal ions affect insulin-degrading enzyme activity

“…This inhibition is even more significant in the case of silver(I) (which is employed instead of copper(I), due to the higher stability of the monovalent form under the experimental conditions), such that the enzyme is completely inactive toward the B20-30 peptide, similar to what has been observed in the case of the Aβ [16][17][18][19][20][21][22][23][24][25][26][27][28] peptide [54], and it is not reported in Fig. 6.…”

“…We carried out enzymatic digestions of B20-30, in the presence or absence of copper(II), aluminum(III) and zinc(II), metals chosen for their important role in neurodegenerative diseases [24,26,66,67]. Moreover, since copper enters the cell as copper(I) through high-affinity plasma membrane copper transporters or low affinity permeases [68] and IDE is mainly cytosolic [1], it is interesting to analyze the effect of both oxidation states on B (20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30) processing. Furthermore, as silver belongs to the same group of copper and silver(I) has the same chemistry of copper(I), we have also performed studies using this ion, in order to have a replication of the copper(I) results without any possible invalidation due to copper(I) oxidation problems (although we worked in a reducing environment, copper(I) could be oxidized to copper(II) during the incubation with the enzyme).…”

“…To assess how metal ions affect the IDE enzymatic activity, steady state kinetics on the B (20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30) peptide in the presence and in the absence of the above mentioned ions were performed. Fig.…”

“…It is also interesting to outline that if we compare these parameters with those obtained for the amyloid peptides Aβ [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16] and Aβ [16][17][18][19][20][21][22][23][24][25][26][27][28] several similarities and some differences emerge [54]. In particular, the overall proteolytic activity toward the main cleavage sites (i.e., Phe 24 [16][17][18][19][20][21][22][23][24][25][26][27][28] ) is somewhat higher in the case of the B20-30 peptide due to a slightly more efficient cleavage rate-limiting step (see Table 2), whereas the substrate affinity of B20-30 appears intermediate between that of Aβ [1][2][3]…”

“…It is often reported that the common feature shared by IDE substrates is the ability to form, under certain physiological conditions, amyloid fibrils [18][19][20][21]. Furthermore, several evidences indicate that the selective binding of IDE's substrates and their unfolding and processing are driven by the unique size, shape and electrostatic potential of the catalytic chamber and by interaction with the exosite [22,23].…”