Dataset Information

Mechanisms of ?-Secretase Activation and Substrate Processing.

ABSTRACT: Amyloid ?-peptide, the principal component of characteristic cerebral plaques of Alzheimer's disease (AD), is produced through intramembrane proteolysis of the amyloid precursor protein (APP) by ?-secretase. Despite the importance in the pathogenesis of AD, the mechanisms of intramembrane proteolysis and substrate processing by ?-secretase remain poorly understood. Here, complementary all-atom simulations using a robust Gaussian accelerated molecular dynamics (GaMD) method and biochemical experiments were combined to investigate substrate processing of wildtype and mutant APP by ?-secretase. The GaMD simulations captured spontaneous activation of ?-secretase, with hydrogen bonded catalytic aspartates and water poised for proteolysis of APP at the ? cleavage site. Furthermore, GaMD simulations revealed that familial AD mutations I45F and T48P enhanced the initial ? cleavage between residues Leu49-Val50, while M51F mutation shifted the ? cleavage site to the amide bond between Thr48-Leu49. Detailed analysis of the GaMD simulations allowed us to identify distinct low-energy conformational states of ?-secretase, different secondary structures of the wildtype and mutant APP substrate, and important active-site subpockets for catalytic function of the enzyme. The simulation findings were highly consistent with experimental analyses of APP proteolytic products using mass spectrometry and Western blotting. Taken together, the GaMD simulations and biochemical experiments have enabled us to elucidate the mechanisms of ?-secretase activation and substrate processing, which should facilitate rational computer-aided drug design targeting this functionally important enzyme.

SUBMITTER: Bhattarai A

PROVIDER: S-EPMC7318072 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Mechanisms of γ-Secretase Activation and Substrate Processing.

Bhattarai Apurba A Devkota Sujan S Bhattarai Sanjay S Wolfe Michael S MS Miao Yinglong Y

ACS central science 20200604 6

Amyloid β-peptide, the principal component of characteristic cerebral plaques of Alzheimer's disease (AD), is produced through intramembrane proteolysis of the amyloid precursor protein (APP) by γ-secretase. Despite the importance in the pathogenesis of AD, the mechanisms of intramembrane proteolysis and substrate processing by γ-secretase remain poorly understood. Here, complementary all-atom simulations using a robust Gaussian accelerated molecular dynamics (GaMD) method and biochemical experi ...[more]

PMID: 32607444

Similar Datasets

Project description:The amyloid β-peptide (Aβ) of Alzheimer's disease (AD) is generated by proteolysis within the transmembrane domain (TMD) of a C-terminal fragment of the amyloid β protein-precursor (APP CTFβ) by the γ-secretase complex. This processing produces Aβ ranging from 38 to 49 residues in length. Evidence suggests that this spectrum of Aβ peptides is the result of successive γ-secretase cleavages, with endoproteolysis first occurring at the ε sites to generate Aβ48 or Aβ49, followed by C-terminal trimming mostly every three residues along two product lines to generate shorter, secreted forms of Aβ: the primary Aβ49-46-43-40 line and a minor Aβ48-45-42-38 line. The major secreted Aβ species are Aβ40 and Aβ42, and an increased proportion of the longer, aggregation-prone Aβ42 compared to Aβ40 is widely thought to be important in AD pathogenesis. We examined TMD substrate determinants of the specificity and efficiency of ε site endoproteolysis and carboxypeptidase trimming of CTFβ by γ-secretase. We determined that the C-terminal negative charge of the intermediate Aβ49 does not play a role in its trimming by γ-secretase. Peptidomimetic probes suggest that γ-secretase has S1', S2', and S3' pockets, through which trimming by tripeptides may be determined. However, deletion of residues around the ε sites demonstrates that a depth of three residues within the TMD is not a determinant of the location of endoproteolytic ε cleavage of CTFβ. We also show that instability of the CTFβ TMD helix near the ε site significantly increases endoproteolysis, and that helical instability near the carboxypeptidase cleavage sites facilitates C-terminal trimming by γ-secretase. In addition, we found that CTFβ dimers are not endoproteolyzed by γ-secretase. These results support a model in which initial interaction of the array of residues along the undimerized single helical TMD of substrates dictates the site of initial ε cleavage and that helix unwinding is essential for both endoproteolysis and carboxypeptidase trimming.

Project description:Neurexins are a large family of neuronal plasma membrane proteins, which function as trans-synaptic receptors during synaptic differentiation. The binding of presynaptic neurexins to postsynaptic partners, such as neuroligins, has been proposed to participate in a signaling pathway that regulates synapse formation/stabilization. The identification of mutations in neurexin genes associated with autism and mental retardation suggests that dysfunction of neurexins may underlie synaptic defects associated with brain disorders. However, the mechanisms that regulate neurexin function at synapses are still unclear. Here, we show that neurexins are proteolytically processed by presenilins (PS), the catalytic components of the γ-secretase complex that mediates the intramembraneous cleavage of several type I membrane proteins. Inhibition of PS/γ-secretase by using pharmacological and genetic approaches induces a drastic accumulation of neurexin C-terminal fragments (CTFs) in cultured rat hippocampal neurons and mouse brain. Neurexin-CTFs accumulate mainly at the presynaptic terminals of PS conditional double knockout (PS cDKO) mice lacking both PS genes in glutamatergic neurons of the forebrain. The fact that loss of PS function enhances neurexin accumulation at glutamatergic terminals mediated by neuroligin-1 suggests that PS regulate the processing of neurexins at glutamatergic synapses. Interestingly, presenilin 1 (PS1) is recruited to glutamatergic terminals mediated by neuroligin-1, thus concentrating PS1 at terminals containing β-neurexins. Furthermore, familial Alzheimer's disease (FAD)-linked PS1 mutations differentially affect β-neurexin-1 processing. Expression of PS1 M146L and PS1 H163R mutants in PS-/- cells rescues the processing of β-neurexin-1, whereas PS1 C410Y and PS1 ΔE9 fail to rescue the processing defect. These results suggest that PS regulate the synaptic function and processing of neurexins at glutamatergic synapses, and that impaired neurexin processing by PS may play a role in FAD.

Dataset Information

Mechanisms of ?-Secretase Activation and Substrate Processing.

Publications

Mechanisms of γ-Secretase Activation and Substrate Processing.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets