Active site geometry stabilization of a presenilin homolog by the lipid bilayer promotes intramembrane proteolysis

Cleavage of membrane proteins in the lipid bilayer by intramembrane proteases is crucial for health and disease. Although different lipid environments can potently modulate their activity, how this is linked to their structural dynamics is unclear. Here we show that the carboxy-peptidase-like activity of the archaeal intramembrane protease PSH, a homolog of the Alzheimer's disease-associated presenilin/γ-secretase is impaired in micelles and promoted in a lipid bilayer. Comparative molecular dynamics simulations revealed that important elements for substrate binding such as transmembrane domain 6a of PSH are more labile in micelles and stabilized in the lipid bilayer. Moreover, consistent with an enhanced interaction of PSH with a transition-state analog inhibitor, the bilayer promoted the formation of the enzyme's catalytic active site geometry. Our data indicate that the lipid environment of an intramembrane protease plays a critical role in structural stabilization and active site arrangement of the enzyme-substrate complex thereby promoting intramembrane proteolysis.

Regulation of neurogenesis and cerebral angiogenesis by cell protein proteolysis products

Brain development is a unique process characterized by mechanisms defined as neuroplasticity (synaptogenesis, synapse elimination, neurogenesis, and cerebral angiogenesis). Numerous neurodevelopmental disorders brain damage, and aging are manifested by neurological deficits that are caused by aberrant neuroplasticity. The presence of stem and progenitor cells in neurogenic niches of the brain is responsible for the formation of new neurons capable of integrating into preexisting synaptic assemblies. The determining factors for the cells within the neurogenic niche are the activity of the vascular scaffold and the availability of active regulatory molecules that establish the optimal microenvironment. It has been found that regulated intramembrane proteolysis plays an important role in the control of neurogenesis in brain neurogenic niches. Molecules generated by the activity of specific proteases can stimulate or suppress the activity of neural stem and progenitor cells, their proliferation and differentiation, migration and integration of newly formed neurons into synaptic networks. Local neoangiogenesis supports the processes of neurogenesis in neurogenic niches, which is guaranteed by the multivalent action of peptides formed from transmembrane proteins. Identification of new molecules regulating the neuroplasticity (neurogenesis and angiogenesis). i. e. enzymes, substrates, and products of intramembrane proteolysis, will ensure the development of protocols for detecting the neuroplasticity markers and targets for efficient pharmacological modulation.

Design and modular assembly of synthetic intramembrane proteolysis receptors for custom gene regulation in therapeutic cells

Synthetic biology has established powerful tools to precisely control cell function. Engineering these systems to meet clinical requirements has enormous medical implications. Here, we adopted a clinically driven design process to build receptors for the autonomous control of therapeutic cells. We examined the function of key domains involved in regulated intramembrane proteolysis and showed that systematic modular engineering can generate a class of receptors we call SyNthetic Intramembrane Proteolysis Receptors (SNIPRs) that have tunable sensing and transcriptional response abilities. We demonstrate the potential transformative utility of the receptor platform by engineering human primary T cells for multi-antigen recognition and production of dosed, bioactive payloads relevant to the treatment of disease. Our design framework enables the development of fully humanized and customizable transcriptional receptors for the programming of therapeutic cells suitable for clinical translation.

Molecular Basis of the Versatile Regulatory Mechanism of HtrA-Type Protease AlgW from Pseudomonas aeruginosa

ABSTRACT AlgW, a membrane-bound periplasmic serine protease belonging to the HtrA protein family, is a key regulator of the regulated intramembrane proteolysis (RIP) pathway and is responsible for transmitting the envelope stress signals in Pseudomonas aeruginosa. The AlgW PDZ domain senses and binds the C-terminal of mis-localized outer membrane proteins (OMPs) or periplasmic protein MucE, leading to catalytic activation of the protease domain. While AlgW is functionally well studied, its exact activation mechanism remains to be elucidated. Here, we show that AlgW is a novel HtrA protease that can be biochemically activated by both peptide and lipid signals. Compared with the corresponding homologue DegS in Escherichia coli, AlgW exhibits a distinct substrate specificity and regulation mechanism. Structural, biochemical, and mutagenic analyses revealed that, by specifically binding to the C-terminal decapeptide of MucE, AlgW could adopt more relaxed conformation and obtain higher activity than with tripeptide activation. We also investigated the regulatory mechanism of the LA loop in AlgW and proved that the unique structural feature of this region was responsible for the distinct enzymatic property of AlgW. These results demonstrate the unique and diverse activation mechanism of AlgW, which P. aeruginosa may utilize to enhance its adaptability to environmental stress. IMPORTANCE HtrA-family proteases are commonly employed to sense the protein folding stress and activate the regulated intramembrane proteolysis (RIP) cascade in Gram-negative bacteria. Here, we reveal the unique dual-signal activation and dynamic regulation properties of AlgW, an HtrA-type protease triggering the AlgU stress-response pathway, which controls alginate production and mucoid conversion in Pseudomonas aeruginosa. The structural and functional data offer insights into the molecular basis underlying the transition of different activation states of AlgW in response to different effectors. Probing these unique features provides an opportunity to correlate the diverse regulation mechanism of AlgW with the high adaptability of P. aeruginosa to environmental changes during infection.

A combination of solid-state NMR and MD simulations reveals the binding mode of a rhomboid protease inhibitor

Intramembrane proteolysis plays a fundamental role in many biological and pathological processes. Intramembrane proteases thus represent promising pharmacological targets, but few selective inhibitors have been identified. This is in contrast...