Because of their abundance, resistance to proteolysis, rapid aggregation and neurotoxicity, N-terminally truncated and, in particular, pyroglutamate (pE)-modified Abeta peptides have been suggested as being important in the initiation of pathological cascades resulting in the development of Alzheimer's disease. We found that the N-terminal pE-formation is catalyzed by glutaminyl cyclase in vivo. Glutaminyl cyclase expression was upregulated in the cortices of individuals with Alzheimer's disease and correlated with the appearance of pE-modified Abeta. Oral application of a glutaminyl cyclase inhibitor resulted in reduced Abeta(3(pE)-42) burden in two different transgenic mouse models of Alzheimer's disease and in a new Drosophila model. Treatment of mice was accompanied by reductions in Abeta(x-40/42), diminished plaque formation and gliosis and improved performance in context memory and spatial learning tests. These observations are consistent with the hypothesis that Abeta(3(pE)-42) acts as a seed for Abeta aggregation by self-aggregation and co-aggregation with Abeta(1-40/42). Therefore, Abeta(3(pE)-40/42) peptides seem to represent Abeta forms with exceptional potency for disturbing neuronal function. The reduction of brain pE-Abeta by inhibition of glutaminyl cyclase offers a new therapeutic option for the treatment of Alzheimer's disease and provides implications for other amyloidoses, such as familial Danish dementia.

Extracellular plaques of β-amyloid (Aβ) and intraneuronal neurofibrillary tangles made from tau are the histopathological signatures of Alzheimer's disease (AD). Plaques comprise Aβ fibrils that assemble from monomeric and oligomeric intermediates, and are prognostic indicators of AD. Despite the significance of plaques to AD, oligomers are considered to be the principal toxic forms of Aβ 1,2 . Interestingly, many adverse responses to Aβ, such as cytotoxicity 3 , microtubule loss 4 , impaired memory and learning 5 , and neuritic degeneration 6 , are greatly amplified by tau expression. N-terminally truncated, pyroglutamylated (pE) forms of Aβ 7,8 are strongly associated with AD, are more toxic than Aβ 1-42 and Aβ , and have been proposed as initiators of AD Users may view, print, copy, download and text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: http://www.nature.com/authors/editorial_policies/license.html#terms * Correspondence: gsb4g@virginia.edu. **Correspondence: Hans-Ulrich.Demuth@probiodrug.de. J.M.N and S.S. contributed equally to the paper.Full Methods and relevant references will be available in the online Supplementary Information accompanying this paper at http:// www.nature.com/nature.Author Contributions: J.M.N. performed most of the biochemical and cell biological experiments; S.S. was the principal force behind the experiments involving hAPP SL /hQC and TBA2.1/tau KO mice, and was aided by B.H.-P., H.C.; A.S. and T.W. fractionated and analyzed human brain extracts; E.S., K.Y. and B.W. performed the peri-hippocampal injection experiments; A.H. and C.G.G. produced and characterized the M64 and M87 antibodies; R.R. and K.R. performed the electrophysiology experiments; A.A., W.J. and S.G. performed and analyzed the immunohistochemical experiments on TBA2.1 and Tau-KO/TBA2.1 mice; G.S.B. and H.-U.D. initiated and directed the project; G.S.B. was the principal writer of the paper; all of the authors participated in the design and analysis of experiments, and in editing of the paper. Fig. 2) to the oligomers. HHS Public AccessAt 5 μM peptide, 5% pE-Aβ aggregated faster than Aβ 3(pE)-42 or Aβ 1-42 alone based on thioflavin T fluorescence shifts 15 ( Supplementary Fig. 3). The OD 450 /OD 490 ratio for Aβ 3(pE)-42 rose and peaked more rapidly than for Aβ 1-42 , but peaked at an ~25% lower level. The fastest rise in the OD 450 /OD 490 ratio was for 5% pE-Aβ, which peaked similarly to Aβ 3(pE)-42 . Aβ 3(pE)-42 , Aβ 1-42 and 5% pE-Aβ thus oligomerized by different pathways.To test whether distinct biological activities were coupled to these oligomerization differences, we compared cytotoxicity of the peptides towards cultured neurons or glia using calcein-AM and fluorescence microscopy 16 . Twelve hours of Aβ 1-42 exposure had little effect on cell viability for wild type (WT) or tau knockout (KO) neurons, or WT glial cells (Fig. 1a). Contrastingly, most WT neurons died and detached from the substrate after exposur...

Oligomerization of amyloid beta (Abeta) peptides is the decisive event in the development of Alzheimer's disease (AD), the most common neurogenerative disorder in developed countries. Recent evidence links this conformation-driven process to primary- and secondary-structure modifications of Abeta. The N and C terminus of deposited Abeta has been shown to possess conspicuous heterogeneity. While the C-terminally longer form of Abeta, i.e., Abeta (42), is considered more amyloidogenic, the role of the N-terminal modifications, e.g., truncation and glutamate cyclization accounting for the majority of the deposited peptides, is less understood. In the present study, we characterized the oligomerization and seeding capacity of pGlu-amyloid peptides using two unrelated techniques based on flow cytometry or flourescence dye binding. Under different conditions and irrespective of the C terminus of Abeta, i.e., Abeta40 or 42, pGlu-modified peptides displayed an up to 250-fold accelerated initial formation of aggregates compared to unmodified Abeta. The accelerated seed formation is accompanied by a change in the oligomerization kinetics because of N-terminal pGlu formation. Furthermore, the formation of mixed aggregates consisting of either pGlu-Abeta (3-42) or ADan or ABri and Abeta (1-42) was investigated by Abeta fluorescence labeling in flow cytometry. The results suggest that pGlu-modified peptides are potential seeding species of aggregate formation in vivo. The data presented here and the abundance of pGlu peptides in amyloidoses, such as FBD and AD, suggest pGlu-amyloid peptides as a species with biophysical characteristics that might be in particular crucial for the initiation of the disease.

N-terminal pyroglutamate (pGlu) formation from glutaminyl precursors is a posttranslational event in the processing of bioactive neuropeptides such as thyrotropin-releasing hormone and neurotensin during their maturation in the secretory pathway. The reaction is facilitated by glutaminyl cyclase (QC), an enzyme highly abundant in mammalian brain. Here, we describe for the ¢rst time that human and papaya QC also catalyze N-terminal glutamate cyclization. Surprisingly, the enzymatic Glu 1 conversion is favored at pH 6.0 while Gln 1 conversion occurs with an optimum at pH 8.0. This unexpected ¢nding might be of importance for deciphering the events leading to deposition of highly toxic pyroglutamyl peptides in amyloidotic diseases. ß 2004 Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.

N-Terminally truncated and pyroglutamate (pGlu) modified amyloid beta (Abeta) peptides are major constituents of amyloid deposits in sporadic and inherited Alzheimer's disease (AD). Formation of pGlu at the N-terminus confers resistance against cleavage by most aminopeptidases, increases toxicity of the peptides, and may seed Abeta aggregate formation. Similarly, the deposited amyloid peptides ABri and ADan, which cause a very similar histopathology in familial British dementia (FBD) and familial Danish dementia (FDD), are N-terminally blocked by pGlu. Triggered by the coincidence of pGlu-modified amyloid peptides and similar pathology in AD, FBD, and FDD, we investigated the impact of N-terminal pGlu on biochemical and biophysical properties of Abeta, ABri, and ADan. N-Terminal pGlu increases the hydrophobicity and changes the pH-dependent solubility profile, rendering the pGlu-modified peptides less soluble in the basic pH range. The pGlu residue increases the aggregation propensity of all amyloid peptides as evidenced by ThT fluorescence assays and dynamic light scattering. The far-UV CD spectroscopic analysis points toward an enhanced beta-sheet structure of the pGlu-Abeta. Importantly, changes in fibril morphology are clearly caused by the N-terminal pGlu, resulting in the formation of short fibers, which are frequently arranged in bundles. The effect of pGlu on the morphology is virtually indistinguishable between ABri, ADan, and Abeta. The data provide evidence for a comparable influence of the pGlu modification on the aggregation process of structurally different amyloid peptides, thus likely contributing to the molecularly distinct neurodegenerative diseases AD, FBD, and FDD. The main driving force for the aggregation is apparently an increase in the hydrophobicity and thus an accelerated seed formation.