HTRA2 Human
Description
Overview and Molecular Characterization of HTRA2 Human
HTRA2 is encoded by the HTRA2 gene located on chromosome 2p13.1 (NCBI: NG_012163.1) and translates into a 49 kDa protein . The mature protein (residues 134–458) contains:
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Serine protease domain: Catalytic triad (H198, D228, S306) essential for proteolytic activity
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PDZ domain: Modulates substrate binding and enzymatic regulation
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N-terminal mitochondrial targeting sequence (residues 1–133): Cleaved during maturation
Protease Activity
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Cleaves misfolded proteins (e.g., amyloid-β, mitochondrial APP)
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Activated through trimerization and PDZ domain rearrangement
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Displays viscosity-dependent biphasic kinetics, suggesting motion-dependent regulation
Chaperone Activity
Apoptotic Regulation
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Caspase-independent: Degrades inhibitor of apoptosis proteins (IAPs) via AVPS motif
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Caspase-dependent: Amplifies mitochondrial death signals under stress
Parkinson’s Disease (PARK13)
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Loss-of-function mutations (e.g., G399S) impair mitochondrial proteostasis, leading to:
Alzheimer’s Disease
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Reduces Aβ42 fibrillization by 68% in vitro through dual mechanisms :
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Proteolytic cleavage of Aβ42 precursors
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Chaperone-mediated inhibition of amyloid aggregation
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Research Findings and Therapeutic Insights
Key Studies
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Substrate Processing Mechanism (PNAS 2022) :
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HTRA2 binds unfolded substrates via PDZ domain and central hydrophobic regions
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Multivalent substrate interactions enhance catalytic efficiency by 4.8×
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Chaperone-Protease Switch (PMC 2007) :
Condition Protease Activity Chaperone Activity Full-length HTRA2 Moderate Low ΔPDZ HTRA2 Absent High -
Allosteric Regulation (Front. Mol. Biosci. 2022) :
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Trimeric architecture enables cooperative activation between protomers
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PDZ-protease crosstalk regulates substrate access to active sites
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Pathogenic Variants and Clinical Significance
The LOVD database documents 34 HTRA2 variants linked to:
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3-Methylglutaconic Aciduria Type VIII (MGCA8): Mitochondrial metabolic disorder
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Early-Onset Parkinsonism (PARK13): 27 unique pathogenic mutations
Notable Mutations
Product Specs
Q&A
Basic Research Questions
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What is the basic structure and function of human HTRA2?
Human HTRA2 is a member of the evolutionarily conserved HtrA family of serine proteases that shows extensive homology to bacterial HtrA genes essential for survival at high temperatures . The protein contains a serine protease domain responsible for its proteolytic activity and a PDZ domain involved in regulatory functions. HTRA2 exists predominantly as a trimeric complex with each monomer containing approximately 458 amino acids. The protein undergoes processing to reveal an N-terminal tetrapeptide (AVPS) motif upon maturation . Functionally, HTRA2 serves dual roles: as a protease that cleaves various substrates including Inhibitor of Apoptosis Proteins (IAPs), and as a chaperone that prevents protein aggregation . The active site contains a catalytic triad with serine 306 being critical for proteolytic activity, as demonstrated by site-directed mutagenesis studies where altering this residue to alanine abolishes autoproteolysis .
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Where is HTRA2 localized in human cells?
Unlike its human paralogs (HtrA1, HtrA3, and HtrA4) that are primarily found in secretory pathways, HTRA2 is predominantly localized to the mitochondrial intermembrane space (IMS) . This distinct localization is directed by an N-terminal mitochondrial targeting sequence. The protein initially anchors to the mitochondrial membrane through a transmembrane domain before undergoing processing to its mature form. Under cellular stress conditions or during apoptotic events, HTRA2 can be released from the mitochondria into the cytosol, where it contributes to cell death pathways by interacting with and cleaving IAPs . Experimental verification of HTRA2's mitochondrial localization typically employs techniques such as subcellular fractionation, immunofluorescence microscopy, and biochemical isolation of mitochondria.
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What experimental methods are commonly used to study HTRA2 activity?
Researchers employ various complementary techniques to study HTRA2's diverse activities:
Functional Aspect Experimental Approach Detection Method Proteolytic activity β-casein cleavage assay Gel electrophoresis, spectroscopic detection Autoproteolysis Site-directed mutagenesis (S306A) Western blot analysis Chaperone function Citrate synthase aggregation assay Light scattering measurements Aβ peptide interaction Electron microscopy, NMR Visualization of fiber formation Temperature-dependent activity Comparative assays at 37°C vs. 45°C Quantification of substrate turnover For cellular studies, researchers typically use cell fractionation to isolate mitochondria, immunoblotting to detect HTRA2 and its substrates, and genetic approaches including knockdown or knockout models to assess functional consequences . Biochemical characterization often involves recombinant protein production, followed by in vitro assays using model substrates under controlled conditions.
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How does HTRA2 contribute to mitochondrial function?
HTRA2 plays a critical role in maintaining mitochondrial homeostasis through several mechanisms:
First, it functions as a quality control protease in the intermembrane space, eliminating damaged or misfolded proteins that could otherwise form toxic aggregates. Second, HTRA2 influences mitochondrial dynamics through its interaction with and processing of OPA1 (Optic Atrophy 1), a key regulator of mitochondrial fusion and cristae morphology . Neural-specific deletion of Htra2 in mice leads to profound mitochondrial abnormalities, including swelling, vesiculation, and fragmentation of cristae, with defective processing of OPA1 specifically resulting in depletion of the L-isoform . These structural abnormalities precede neurodegeneration, indicating that mitochondrial dysfunction is a primary consequence of HTRA2 deficiency rather than a secondary effect. Without functional HTRA2, cells accumulate dysfunctional mitochondria, which contributes to neuronal death and premature mortality in animal models .
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What is the role of HTRA2 in cellular stress response?
HTRA2 serves as a critical component of the cellular stress response pathway, particularly in the context of mitochondrial proteostasis. The protein is upregulated in mammalian cells in response to various stressors, including heat shock and tunicamycin treatment, which induces ER stress . This upregulation pattern is consistent with its bacterial homologs that are essential for survival at elevated temperatures. Under stress conditions, HTRA2 helps prevent the accumulation of misfolded or denatured proteins in the mitochondrial intermembrane space through both its proteolytic and chaperone activities . The temperature sensitivity of HTRA2 is evidenced by increased β-casein turnover when assay temperature is raised from 37°C to 45°C, suggesting enhanced proteolytic function at higher temperatures . This biochemical property, combined with its nuclear localization and stress-induced expression, positions HTRA2 as a key player in mitochondrial stress signaling and cellular adaptation to proteotoxic stress.
Product Science Overview
HTRA2 is synthesized as a precursor protein and is processed to its mature form within the mitochondria. The mature protein contains a serine protease domain and a PDZ domain, which are crucial for its proteolytic activity and substrate recognition, respectively . The protein is involved in the degradation of misfolded or damaged proteins within the mitochondria, thereby maintaining mitochondrial function and integrity .
One of the key functions of HTRA2 is its role in apoptosis. Under stress conditions, HTRA2 is released from the mitochondria into the cytosol, where it interacts with and cleaves various substrates, including inhibitor of apoptosis proteins (IAPs). This cleavage leads to the activation of caspases, which are the executioners of apoptosis . Thus, HTRA2 acts as a pro-apoptotic factor, promoting cell death in response to cellular stress.
Mutations in the HTRA2 gene have been associated with several diseases, including Parkinson’s disease and 3-methylglutaconic aciduria, type VIII . In Parkinson’s disease, mutations in HTRA2 are thought to impair its proteolytic activity, leading to the accumulation of damaged proteins and mitochondrial dysfunction, which contribute to the degeneration of dopaminergic neurons .
Recombinant HTRA2 is produced using various expression systems, such as E. coli, to study its structure, function, and role in disease. The recombinant protein is often used in biochemical assays to investigate its proteolytic activity and interactions with other proteins . For instance, recombinant HTRA2 has been shown to cleave beta-casein, a model substrate, demonstrating its protease activity .
HTRA2 is a subject of extensive research due to its dual role in maintaining mitochondrial homeostasis and regulating apoptosis. Studies have shown that HTRA2 is involved in the response to cellular stress and the regulation of mitochondrial dynamics . Additionally, HTRA2 is being investigated as a potential therapeutic target for diseases associated with mitochondrial dysfunction and impaired apoptosis, such as neurodegenerative diseases and cancer .
