Several single-span membrane proteins are cleaved within their transmembrane domains (TMDs) by intramembrane-cleaving proteases, although the structure of the active site executing intramembrane cleavage remains unknown. Here we use the substituted cysteine accessibility method to examine the structure of presenilin-1, a catalytic subunit of gamma-secretase, involved in amyloid beta protein generation in Alzheimer's disease and Notch signaling. We show that TMD6 and TMD7 of presenilin-1 contribute to the formation of a hydrophilic pore within the membrane. Residues at the luminal portion of TMD6 are predicted to form a subsite for substrate or inhibitor binding on the alpha-helix facing a hydrophilic milieu, whereas those around the GxGD catalytic motif within TMD7 are highly water accessible, suggesting formation of a hydrophilic structure within the pore. Collectively, our data suggest that the active site of gamma-secretase resides in a catalytic pore filled with water within the lipid bilayer and is tapered around the catalytic aspartates.
"Eight kDa N-terminal fragment of PS1 NTF generated by thrombin cleavage of PS1-Th1 after Fen-B crosslinking was specifically precipitated in a similar fashion to that by phenylpiperidine-type photoprobes, suggesting that Fen-B also targets to the most N-terminal region of PS1, including TMD1 (Figure
2B and C). We showed that the cytosolic side of TMD1 participates in the catalytic hydrophilic pore
[7,11]. To analyze the relationship between the fenofibrate binding site and the catalytic site within TMD1, we employed the cross-competition analysis in Fen-B labeling using different classes of compounds. "
[Show abstract][Hide abstract] ABSTRACT: Amyloid-beta peptide ending at 42nd residue (Abeta42) is believed as a pathogenic peptide for Alzheimer disease. Although gamma-secretase is a responsible protease to generate Abeta through a processive cleavage, the proteolytic mechanism of gamma-secretase at molecular level is poorly understood.
We found that the transmembrane domain (TMD) 1 of presenilin (PS) 1, a catalytic subunit for the gamma-secretase, as a key modulatory domain for Abeta42 production. Abeta42-lowering and -raising gamma-secretase modulators (GSMs) directly targeted TMD1 of PS1 and affected its structure. A point mutation in TMD1 caused an aberrant secretion of longer Abeta species including Abeta45 that are the precursor of Abeta42. We further found that the helical surface of TMD1 is involved in the binding of Abeta45/48 and that the binding was altered by GSMs as well as TMD1 mutation.
Binding between PS1 TMD1 and longer Abeta is critical for Abeta42 production.
"Electronic microscopic analysis and single particle imaging revealed the existence of intramembrane water-accessible cylindrical chamber in gamma-secretase with a low-density cavity from extracellular side [78,79]. Parallel substituted cysteine accessible method (SCAM) and cross-link experiment confirmed that TM6, TM7 and TM9 of PS formed the intramembrane chamber with two catalytic aspartates residing oppositely on TM6 and TM7, respectively [80-84]. The constitutive autoendoproteolysis of PS removes the inhibitory allosteric effect of the large hydrophobic loop from the catalytic chamber structure in PS [64,65]. "
[Show abstract][Hide abstract] ABSTRACT: Presenilins (PSs) are the catalytic core of gamma-secretase complex. However, the mechanism of FAD-associated PS mutations in AD pathogenesis still remains elusive. Here we review the general biology and mechanism of gamma-secretase and focus on the catalytic components -- presenilins and their biological functions and contributions to the AD pathogenesis. The functions of presenilins are divided into gamma-secretase dependent and gamma-secretase independent ones. The gamma-secretase dependent functions of presenilins are exemplified by the sequential cleavages in the processing of APP and Notch; the gamma-secretase independent functions of presenilins include stabilizing beta-catenin in Wnt signaling pathway, regulating calcium homeostasis and their interaction with synaptic transmission.
"In 2006 the hydrophilic cavity, formed by TMD6 and TMD7, and containing YD and GXGD motifs, was discovered. This cavity or pore was suggested as a catalytic site (Sato el al. 2006; Tolia et al. 2006). Two important points have been recently added to this observation through crystallographic analysis of the archeal protein MCMJR1, a PS homolog. "
[Show abstract][Hide abstract] ABSTRACT: Presenilin is a central, catalytic component of the γ-secretase complex which conducts intramembrane cleavage of various protein substrates. Although identified and mainly studied through its role in the development of amyloid plaques in Alzheimer disease, γ-secretase has many other important functions. The complex seems to be evolutionary conserved throughout the Metazoa, but recent findings in plants and Dictyostelium discoideum as well as in archeons suggest that its evolution and functions might be much more diversified than previously expected. In this review, a selective survey of the multitude of functions of presenilins and the γ-secretase complex is presented. Following a brief overview of γ-secretase structure, assembly and maturation, three functional aspects are analyzed: (1) the role of γ-secretase in autophagy and phagocytosis; (2) involvement of the complex in signaling related to endocytosis; and (3) control of calcium fluxes by presenilins.
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