Hypochlorite-induced oxidation promotes aggregation and reduces toxicity of amyloid beta 1-42.

Exacerbated hypochlorite (OCl) production is linked to neurodegenerative processes, but there is growing evidence that lower levels of hypochlorite activity are important to protein homeostasis. In this study we characterise the effects of hypochlorite on the aggregation and toxicity of amyloid beta peptide 1-42 (Aβ), a major component of amyloid plaques that form in the brain in Alzheimer's disease. Our results demonstrate that treatment with hypochlorite promotes the formation of Aβ assemblies ≥100 kDa that have reduced surface exposed hydrophobicity compared to the untreated peptide.

A Novel Role for Plasminogen Activator Inhibitor Type-2 as a Hypochlorite-Resistant Serine Protease Inhibitor and Holdase Chaperone.

Plasminogen activator inhibitor type-2 (PAI-2), a member of the serpin family, is dramatically upregulated during pregnancy and in response to inflammation. Although PAI-2 exists in glycosylated and non-glycosylated forms in vivo, the majority of in vitro studies of PAI-2 have exclusively involved the intracellular non-glycosylated form. This study shows that exposure to inflammation-associated hypochlorite induces the oligomerisation of PAI-2 via a mechanism involving dityrosine formation.

Hypochlorite-induced aggregation of fibrinogen underlies a novel antioxidant role in blood plasma.

Fibrinogen, a major constituent of blood plasma, is highly susceptible to reaction with biological oxidants. It has been proposed that fibrinogen plays a role in antioxidant defence, but oxidation of fibrinogen is also known to disrupt normal blood clotting and is implicated in the pathology of atherosclerosis. In the present study, we show that the biological oxidant hypochlorite promotes the formation of soluble high molecular weight fibrinogen assemblies ≥40 × 10 Da, that do not accumulate when fibrinogen is induced to aggregate by other stresses such as heating or hydroxyl-mediated damage in vitro.

Alpha-2-Macroglobulin, a Hypochlorite-Regulated Chaperone and Immune System Modulator.

Alpha-macroglobulins are ancient proteins that include monomeric, dimeric, and tetrameric family members. In humans, and many other mammals, the predominant alpha-macroglobulin is alpha-2-macroglobulin ( M), a tetrameric protein that is constitutively abundant in biological fluids (e.g.

Human pregnancy zone protein stabilizes misfolded proteins including preeclampsia- and Alzheimer's-associated amyloid beta peptide.

Protein misfolding underlies the pathology of a large number of human disorders, many of which are age-related. An exception to this is preeclampsia, a leading cause of pregnancy-associated morbidity and mortality in which misfolded proteins accumulate in body fluids and the placenta. We demonstrate that pregnancy zone protein (PZP), which is dramatically elevated in maternal plasma during pregnancy, efficiently inhibits in vitro the aggregation of misfolded proteins, including the amyloid beta peptide (Aβ) that is implicated in preeclampsia as well as with Alzheimer's disease.

Amorphous protein aggregates stimulate plasminogen activation, leading to release of cytotoxic fragments that are clients for extracellular chaperones.

The misfolding of proteins and their accumulation in extracellular tissue compartments as insoluble amyloid or amorphous protein aggregates are a hallmark feature of many debilitating protein deposition diseases such as Alzheimer's disease, prion diseases, and type II diabetes. The plasminogen activation system is best known as an extracellular fibrinolytic system but was previously reported to also be capable of degrading amyloid fibrils. Here we show that amorphous protein aggregates interact with tissue-type plasminogen activator and plasminogen, via an exposed lysine-dependent mechanism, to efficiently generate plasmin.

PZP and PAI-2: Structurally-diverse, functionally similar pregnancy proteins?

Pregnancy zone protein (PZP) and plasminogen activator inhibitor type 2 (PAI-2) are two multifunctional proteins that are elevated in normal pregnancy and numerous other inflammatory states. Both proteins were originally identified as protease inhibitors, but current evidence supports the notion that they may also function as modulators of T-helper cells and/or extracellular chaperones. Exacerbated inflammation, fibrinolytic disturbances and misfolded proteins are all implicated in the pathology of preeclampsia, a leading cause of maternal and foetal mortality and morbidity.

Alpha-2-Macroglobulin Is Acutely Sensitive to Freezing and Lyophilization: Implications for Structural and Functional Studies.

Alpha-2-macroglobulin is an abundant secreted protein that is of particular interest because of its diverse ligand binding profile and multifunctional nature, which includes roles as a protease inhibitor and as a molecular chaperone. The activities of alpha-2-macroglobulin are typically dependent on whether its conformation is native or transformed (i.e.

Hypochlorite-induced structural modifications enhance the chaperone activity of human α2-macroglobulin.

Hypochlorite, an oxidant generated in vivo by the innate immune system, kills invading pathogens largely by inducing the misfolding of microbial proteins. Concomitantly, the nonspecific activity of hypochlorite also damages host proteins, and the accumulation of damaged (misfolded) proteins is implicated in the pathology of a variety of debilitating human disorders (e.g.

Protease-activated alpha-2-macroglobulin can inhibit amyloid formation via two distinct mechanisms.

α(2)-Macroglobulin (α(2)M) is an extracellular chaperone that inhibits amorphous and fibrillar protein aggregation. The reaction of α(2)M with proteases results in an 'activated' conformation, where the proteases become covalently-linked within the interior of a cage-like structure formed by α(2)M. This study investigates, the effect of activation on the ability of α(2)M to inhibit amyloid formation by Aβ(1-42) and I59T human lysozyme and shows that protease-activated α(2)M can act via two distinct mechanisms: (i) by trapping proteases that remain able to degrade polypeptide chains and (ii) by a chaperone action that prevents misfolded clients from continuing along the amyloid forming pathway.

Extracellular chaperones and proteostasis.

There exists a family of currently untreatable, serious human diseases that arise from the inappropriate misfolding and aggregation of extracellular proteins. At present our understanding of mechanisms that operate to maintain proteostasis in extracellular body fluids is limited, but it has significantly advanced with the discovery of a small but growing family of constitutively secreted extracellular chaperones. The available evidence strongly suggests that these chaperones act as both sensors and disposal mediators of misfolded proteins in extracellular fluids, thereby normally protecting us from disease pathologies.

Acute phase proteins are major clients for the chaperone action of α₂-macroglobulin in human plasma.

Extracellular protein misfolding is implicated in many age-related diseases including Alzheimer's disease, macular degeneration and arthritis. In this study, putative endogenous clients for the chaperone activity of α₂-macroglobulin (α₂M) were identified after human plasma was subjected to physiologically relevant sheer stress at 37 °C for 10 days. Western blot analysis showed that four major acute phase proteins: ceruloplasmin, fibrinogen, α₁-acid glycoprotein and complement component 3, preferentially co-purified with α₂M after plasma was stressed.

Roles of extracellular chaperones in amyloidosis.

Extracellular protein misfolding and aggregation underlie many of the most serious amyloidoses including Alzheimer's disease, spongiform encephalopathies and type II diabetes. Despite this, protein homeostasis (proteostasis) research has largely focussed on characterising systems that function to monitor protein conformation and concentration within cells. We are now starting to identify elements of corresponding systems, including an expanding family of secreted chaperones, which exist in the extracellular space.

Extracellular chaperones.

The maintenance of the levels and correct folding state of proteins (proteostasis) is a fundamental prerequisite for life. Life has evolved complex mechanisms to maintain proteostasis and many of these that operate inside cells are now well understood. The same cannot yet be said of corresponding processes in extracellular fluids of the human body, where inappropriate protein aggregation is known to underpin many serious diseases such as Alzheimer's disease, type II diabetes and prion diseases.

Extracellular Chaperones.

The maintenance of the levels and correct folding state of proteins (proteostasis) is a fundamental prerequisite for life. Life has evolved complex mechanisms to maintain proteostasis and many of these that operate inside cells are now well understood. The same cannot yet be said of corresponding processes in extracellular fluids of the human body, where inappropriate protein aggregation is known to underpin many serious diseases such as Alzheimer's disease, type II diabetes and prion diseases.

Clusterin facilitates in vivo clearance of extracellular misfolded proteins.

The extracellular deposition of misfolded proteins is a characteristic of many debilitating age-related disorders. However, little is known about the specific mechanisms that act to suppress this process in vivo. Clusterin (CLU) is an extracellular chaperone that forms stable and soluble complexes with misfolded client proteins.

Identification of human plasma proteins as major clients for the extracellular chaperone clusterin.

Clusterin (CLU) is an extracellular chaperone that is likely to play an important role in protein folding quality control. This study identified three deposition disease-associated proteins as major plasma clients for clusterin by studying CLU-client complexes formed in response to physiologically relevant stress (shear stress, approximately 36 dynes/cm(2) at 37 degrees C). Analysis of plasma samples by size exclusion chromatography indicated that (i) relative to control plasma, stressed plasma contained proportionally more soluble protein species of high molecular weight, and (ii) high molecular weight species were far more abundant when proteins purified by anti-CLU immunoaffinity chromatography from stressed plasma were compared with those purified from control plasma.

Structural characterization of clusterin-chaperone client protein complexes.

Clusterin (CLU) is a potent extracellular chaperone that inhibits protein aggregation and precipitation otherwise caused by physical or chemical stresses (e.g. heat, reduction).

Quality control of protein folding in extracellular space.

The pathologies of many serious human diseases are thought to develop from the effects of intra- or extracellular aggregates of non-native proteins. Inside cells, chaperone and protease systems regulate protein folding; however, little is known about any corresponding mechanisms that operate extracellularly. The identification of these mechanisms is important for the development of new disease therapies.