Structure of nicastrin unveils secrets of γ-secretase.

Despite afflicting tens of millions worldwide, Alzheimer’s disease (AD) remains a devastating brain-destroying malady with no effective treatment or cure. A central hallmark of AD is the deposition of amyloid plaques in the brain. These plaques are primarily composed of the amyloid β-peptide (Aβ), a byproduct of proteolytic processing of the amyloid precursor protein (APP) within its transmembrane domain by γ-secretase. Mutations in both APP and presenilin, the catalytic component of γ-secretase, are associated with dominant early-onset AD, highlighting their central importance in pathogenesis (1). Gaining a detailed mechanistic understanding of how γ-secretase processes APP—and its other substrates, such as Notch receptors— is critically important for biology and medicine (2). Despite a wealth of biochemical studies, key details of how γ-secretase functions have remained elusive because of a lack of high-resolution structural data. In PNAS, Xie et al. present a crystal structure of nicastrin, the first atomic-resolution structure of a component of a γ-secretase complex (3). The membrane-embedded γ-secretase complex is comprised of four proteins that are necessary and sufficient for its activity: Presenilin, Pen-2, Aph-1, and nicastrin (4). Presenilin, an aspartyl protease, contains catalytic aspartates on transmembrane domains (TMDs) 6 and 7 of its nine TMD helices. Upon assembly of the four components within the endoplasmic reticulum, presenilin undergoes autoproteolysis between TMDs 6 and 7 to form catalytically active γ-secretase. Pen2 has two TMDs and is thought to be important for triggering presenilin self-cleavage (5, 6). Little is known about the biochemical function of the sevenTMD Aph-1 other than that it is required for complex formation and full maturation of γ-secretase, although a recent study suggests a role in determining the length of Aβ peptide proteolytic products (7). The fourth component of the complex, nicastrin, is a single-TMD protein with a large ectodomain. γ-Secretase is a member of a broader family of intramembrane-cleaving proteases, which include the site 2 protease (S2P) family of metalloproteases and the rhomboid family of serine proteases (8). Unlike S2P and rhomboid, whose high-resolution structures were solved some years ago, detailed structural information of γ-secretase has been slow to develop. Until recently, the only structural information known about γ-secretase has been gleaned from low-resolution (∼12 Å at best) electron microscopy (EM) studies. With the advent of advanced cryo-EM techniques and equipment, a much more detailed structure of the complex was recently obtained (9). Although the resolution of this structure is still too poor to see atomic details, the overall architecture of the complex was visualized for the first time. Individual TMDs could be seen, although not assigned with certainty to each component of the complex. The most well-resolved portion of the complex was the ectodomain of nicastrin, although all of the glycosylation sites and several long segments of amino acids were not observed, presumably because of their intrinsic flexibility. At 709 amino acids, nicastrin is the largest component of γ-secretase, with the majority of its mass located to its large, heavily glycosylated ectodomain. Although a controversial hypothesis (10), the nicastrin ectodomain has been proposed to bind to the free N terminus of ectodomain-shed substrates of γ-secretase, thereby acting as substrate receptor for the enzyme (11, 12). The mechanism by which this may occur has remained obscure, in part because of a lack of structural information. Xie et al. (3) solved the crystal structure of nicastrin from the amoeboid eukaryote Dictyostelium purpureum, an organism with a complete and functional γ-secretase complex (13). The 1.95 Å-resolution structure (Fig. 1A) reveals a bilobed protein with one large and one small lobe. Five glycosylation sites and six stabilizing disulfide bridges were seen that were not observed in the recent cryo-EM structure of the complex. The two lobes are joined by a collection of hydrophobic interactions at the center of a half circle of hydrogen bonds. With this more complete structure, a structural homology search revealed that the ectodomain of nicastrin most closely resembles that of a bacterial aminopeptidase. Nicastrin, however, lacks the key zinc-binding amino Fig. 1. (A) The crystal structure of nicastrin from the organism D. purpureum at a resolution of 1.95 Å. The bilobed protein is structurally homologous to a bacterial aminopeptidase, although nicastrin itself lacks proteolytic activity. A loop (yellow) extends from the small lobe (red) to cover the putative substrate-binding pocket on the large lobe (blue). Using this structure to improve the cryoEM model of the human γ-secretase complex revealed that this pocket is oriented above the opening of the horseshoe-shaped arrangement of γ-secretase TMDs (blue and green ovals). (B) New speculative arrangement of the TMDs of γ-secretase components within the lipid bilayer. Nicastrin, Aph-1, and presenilin CTF are located to the thick end of the horseshoe shaped structure determined by cryo-EM. Pen2 and presenilin NTF are located to the thin end in this model.