Proteolytic cleavage of Ser52Pro variant transthyretin triggers its amyloid fibrillogenesis

Mangione, Palma; Porcari, Riccardo; Gillmore, Julian D.; Pucci, Piero; Monti, Maria; Porcari, Mattia; Giorgetti, Sofia; Marchese, Loredana; Raimondi, Sara; Serpell, Louise C.; Chen, Wenjie; Relini, Annalisa; Marcoux, Julien; Clatworthy, Innes; Taylor, Graham W.; Tennent, Glenys A.; Robinson, Carol V.; Hawkins, Philip N.; Stoppini, Monica; Wood, Stephen P.; Pepys, Mark B.; Bellotti, Vittorio

doi:10.1073/pnas.1317488111

Cited by 105 publications

(164 citation statements)

References 26 publications

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“…Previous in vitro studies [21,22] have provided evidence that RBP4 can extend the aggregation kinetics of thermally-denatured TTR, implying that RBP4 stabilizes the native conformation of TTR. Based on these findings, we theorized that TTR found in amyloid deposits would no longer be bound to its natural and stabilizing binding partner, RBP4.…”

Section: Discussionmentioning

confidence: 99%

Retinol-binding protein 4 in transthyretin-associated forms of cardiac amyloidosis: differences in the pathobiologies of mutant (ATTRm) and wild-type (ATTRwt) diseases

Koch¹,

Chiswick²,

Cui³

et al. 2017

Clin Diagn Pathol

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The role of retinol-binding protein 4 (RBP4), a natural binding partner of plasma circulating transthyretin (TTR), in TTR-associated cardiomyopathies is unknown. RBP4 is a small (21 kDa) protein that normally functions as a transporter for all-trans retinol (Vitamin A) and travels in the bloodstream as a ternary complex bound to TTR. Previous in vitro studies have demonstrated that RBP4 stabilizes native (tetrameric) TTR and prevents its disassembly into monomers, the key step in TTR amyloid fibril formation. Though increased serum concentrations of RBP4 have been reported in non-amyloidotic cardiomyopathies, there is little information about circulating levels in the inherited (ATTRm) and acquired (ATTRwt) forms of TTR-associated cardiac amyloid disease. Our study objectives were to investigate TTR amyloid-infiltrated cardiac tissue for the presence of RBP4 and to compare serum levels of the protein in ATTRm, ATTRwt and controls. We hypothesized that there would be histological and serological differences in RBP4 between ATTR patient and control samples. In the present study, we demonstrate that RBP4 is highly abundant in ATTRm cardiac tissue surrounding amyloid-infiltrated regions and, to a lesser extent, in ATTRwt specimens. Serum levels of RBP4 are significantly lower in ATTRm compared to ATTRwt (p < 0.0001) and healthy controls (p < 0.0011), with significant correlation between circulating RBP4 concentration and cardiac troponin-I (c-TnI) in our ATTRwt cohort. These data suggest, for the first time, that the pathobiologies of ATTRm and ATTRwt are dissimilar, and provide support for RBP4 as a serum biomarker of amyloid cardiomyopathy in ATTRm.

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Section: Discussionmentioning

confidence: 99%

Retinol-binding protein 4 in transthyretin-associated forms of cardiac amyloidosis: differences in the pathobiologies of mutant (ATTRm) and wild-type (ATTRwt) diseases

Koch¹,

Chiswick²,

Cui³

et al. 2017

Clin Diagn Pathol

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“…The biosynthesis, the various structural domains, cleavage sites and a few selected key mutations in hTTR are shown schematically in Figure 1. The above single monomeric chain forms a strong tetrameric complex (Calculated MW ~55,044 Da) that exhibits its physiological and biological activity [33][34][35]. A recent study with hTTR wild type as well as its key physiological mutant variants suggested that each monomeric form of hTTR is proteolytically cleaved likely by a trypsin like enzyme and this site may be located not far from 50 amino acids from the N-terminal end.…”

Section: Peptide Designmentioning

confidence: 99%

“…A more recent study revealed that for both wild type and variant hTTR, a significantly high level of hTTR (49-127) fragment was found to be present in ex vivo fibril aggregates along with only a small quantity of full length proteins [33]. Moreover the above hTTR sequence domain has also been reported to be the major constitutent of wild type and hTTR variants derived amyloid fibrils linked to cardiomyopathy [34].…”

Section: Introductionmentioning

confidence: 99%

“…However, only a selected number of mutations in hTTR have been strongly linked to high pathogenic severity of amyloidosis. It is suggested that such mutations in hTTR lead to instability of its tetrameric structure which ultimately fold apart as monomer that then self-aggregates into fibril structures promoted by its proteolysis [32,33]. It is now well established that pathogenic severity depends on hTTR aggregation rate and it is most aggressive for the rare L 55 P mutation compared to the more common V 30 M mutation [35].…”

Section: Introductionmentioning

confidence: 99%

“…A number of these mutations are depicted in Figure 1. Lately considerable interest has developed on S 52 P mutation that is considered as physiologically most relevant for increased hTTR aggregation and pathogenesis [33]. The effect of this mutation on hTTR proteolysis is also investigated in this study using synthetic peptides and the result was compared with those from wild type and other mutant hTTR peptides.…”

Section: Introductionmentioning

confidence: 99%

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Proteolytic truncation of human transthyretin linked to amyloidosis is mediated by a trypsin like enzyme: In vitro demonstration using model peptides

Saad¹,

Lu²,

Koya³

et al. 2016

Bio Chem Comp

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Background: A number of human conditions collectively known as amyloidosis were found to be associated with self-aggregation of a specific group of proteins. This fibril like protein aggregates deposit in different tissues including the central nervous system leading to amyloid polyneuropathies (AP), systemic senile amyloidosis (SSA), amyloidotic cardiomyopathy (AC) and carpal tunnel syndrome (CTS). The most common protein in this group is human transthyretin (hTTR). Similar to beta amyloid peptide of Alzheimer's Disease, hTTR forms amyloid type fibrils following its proteolytic cleavage at the C-(carboxy) terminus leading to aggregation. hTTR is normally present in blood where it binds with thyroid hormone thyroxine T4 and Retinol Binding Protein-Vitamin A complex and mediates their transport. It is also present in lower amount in cerebrospinal fluid as a major carrier of thyroxine hormone. Deposition of hTTR fibril aggregates is responsible for SSA usually observed in older population with age >60 years which is regulated by its mutant forms. The exact mechanism of this disorder is still unclear. Studies confirmed that C-terminally truncated hTTR undergoes rapid self-aggregation leading to amyloidosis. The site of this cleavage and the protease involved has not been fully characterized although it is proposed to be at Lys 48 ↓Thr 49 residues.Objective: The major goal of this study is to confirm that hTTR is cleaved by a trypsin like enzyme at the position indicated by vertical arrow Ala-Ser-Gly-Lys 48 ↓Thr 49 -Ser-Glu-Ser and that this cleavage can be blocked by a trypsin-inhibitor. Methods:A 20 mer peptide comprising the wild type sequence from residue (41-60) of hTTR was synthesized, purified by RP-HPLC and fully characterized by mass spectrometry. Three additional mutant peptides namely Lys These results are consistent with those reported for the physiological full length hTTR proteins. Moreover the above cleavage can be blocked efficiently by a trypsin inhibitor (dimethyl formamide), reported for the first time in this study. Energy minimised 3D modeling study revealed that Lys 48 /Ala mutant exhibits a unique structural conformation compared to the corresponding wild type control peptide suggesting its potential role in regulating proteolytic cleavage. Overall our data confirmed the key role of a trypsin like enzyme in hTTR proteolysis leading to its truncation which facilitates its self-aggregation leading to amyloidosis. The study also highlighted a potential therapeutic strategy for hTTR associated amyloidosis by targeting this enzyme with specific inhibitors.

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Cardiac Amyloidosis Due to Transthyretin Protein

Ruberg,

Maurer

2024

JAMA

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ImportanceSystemic amyloidosis from transthyretin (ATTR) protein is the most common type of amyloidosis that causes cardiomyopathy.ObservationsTransthyretin (TTR) protein transports thyroxine (thyroid hormone) and retinol (vitamin A) and is synthesized predominantly by the liver. When the TTR protein misfolds, it can form amyloid fibrils that deposit in the heart causing heart failure, heart conduction block, or arrhythmia such as atrial fibrillation. The biological processes by which amyloid fibrils form are incompletely understood but are associated with aging and, in some patients, affected by inherited variants in the TTR genetic sequence. ATTR amyloidosis results from misfolded TTR protein deposition. ATTR can occur in association with normal TTR genetic sequence (wild-type ATTR) or with abnormal TTR genetic sequence (variant ATTR). Wild-type ATTR primarily manifests as cardiomyopathy while ATTR due to a genetic variant manifests as cardiomyopathy and/or polyneuropathy. Approximately 50 000 to 150 000 people in the US have heart failure due to ATTR amyloidosis. Without treatment, heart failure due to ATTR amyloidosis is associated with a median survival of approximately 5 years. More than 130 different inherited genetic variants in TTR exist. The most common genetic variant is Val122Ile (pV142I), an allele with an origin in West African countries, that is present in 3.4% of African American individuals in the US or approximately 1.5 million persons. The diagnosis can be made using serum free light chain assay and immunofixation electrophoresis to exclude light chain amyloidosis combined with cardiac nuclear scintigraphy to detect radiotracer uptake in a pattern consistent with amyloidosis. Loop diuretics, such as furosemide, torsemide, and bumetanide, are the primary treatment for fluid overload and symptomatic relief of patients with ATTR heart failure. An ATTR-directed therapy that inhibited misfolding of the TTR protein (tafamidis, a protein stabilizer), compared with placebo, reduced mortality from 42.9% to 29.5%, reduced hospitalizations from 0.7/year to 0.48/year, and was most effective when administered early in disease course.Conclusions and RelevanceATTR amyloidosis causes cardiomyopathy in up to approximately 150 000 people in the US and tafamidis is the only currently approved therapy. Tafamidis slowed progression of ATTR amyloidosis and improved survival and prevented hospitalization, compared with placebo, in people with ATTR-associated cardiomyopathy.

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Proteolytic cleavage of Ser52Pro variant transthyretin triggers its amyloid fibrillogenesis

Cited by 105 publications

References 26 publications

Retinol-binding protein 4 in transthyretin-associated forms of cardiac amyloidosis: differences in the pathobiologies of mutant (ATTRm) and wild-type (ATTRwt) diseases

Retinol-binding protein 4 in transthyretin-associated forms of cardiac amyloidosis: differences in the pathobiologies of mutant (ATTRm) and wild-type (ATTRwt) diseases

Proteolytic truncation of human transthyretin linked to amyloidosis is mediated by a trypsin like enzyme: In vitro demonstration using model peptides

Cardiac Amyloidosis Due to Transthyretin Protein

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