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Novel Zn2+-binding sites in human transthyretin: implications for amyloidogenesis and retinol-binding protein recognition.

ABSTRACT: Human transthyretin (TTR) is a homotetrameric protein involved in several amyloidoses. Zn(2+) enhances TTR aggregation in vitro, and is a component of ex vivo TTR amyloid fibrils. We report the first crystal structure of human TTR in complex with Zn(2+) at pH 4.6-7.5. All four structures reveal three tetra-coordinated Zn(2+)-binding sites (ZBS 1-3) per monomer, plus a fourth site (ZBS 4) involving amino acid residues from a symmetry-related tetramer that is not visible in solution by NMR. Zn(2+) binding perturbs loop E-?-helix-loop F, the region involved in holo-retinol-binding protein (holo-RBP) recognition, mainly at acidic pH; TTR affinity for holo-RBP decreases ?5-fold in the presence of Zn(2+). Interestingly, this same region is disrupted in the crystal structure of the amyloidogenic intermediate of TTR formed at acidic pH in the absence of Zn(2+). HNCO and HNCA experiments performed in solution at pH 7.5 revealed that upon Zn(2+) binding, although the ?-helix persists, there are perturbations in the resonances of the residues that flank this region, suggesting an increase in structural flexibility. While stability of the monomer of TTR decreases in the presence of Zn(2+), which is consistent with the tertiary structural perturbation provoked by Zn(2+) binding, tetramer stability is only marginally affected by Zn(2+). These data highlight structural and functional roles of Zn(2+) in TTR-related amyloidoses, as well as in holo-RBP recognition and vitamin A homeostasis.

SUBMITTER: Palmieri Lde C

PROVIDER: S-EPMC2951245 | biostudies-literature | 2010 Oct

REPOSITORIES: biostudies-literature

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Novel Zn2+-binding sites in human transthyretin: implications for amyloidogenesis and retinol-binding protein recognition.

Palmieri Leonardo de C Lde C Lima Luis Mauricio T R LM Freire Juliana B B JB Bleicher Lucas L Polikarpov Igor I Almeida Fabio C L FC Foguel Debora D

The Journal of biological chemistry 20100720 41

Human transthyretin (TTR) is a homotetrameric protein involved in several amyloidoses. Zn(2+) enhances TTR aggregation in vitro, and is a component of ex vivo TTR amyloid fibrils. We report the first crystal structure of human TTR in complex with Zn(2+) at pH 4.6-7.5. All four structures reveal three tetra-coordinated Zn(2+)-binding sites (ZBS 1-3) per monomer, plus a fourth site (ZBS 4) involving amino acid residues from a symmetry-related tetramer that is not visible in solution by NMR. Zn(2+) ...[more]

PMID: 20659897

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Project description:Importance:Transthyretin cardiac amyloidosis (ATTR) is an underrecognized cause of heart failure (HF) in older individuals, owing in part to difficulty in diagnosis. ATTR can result from substitution of valine for isoleucine at codon 122 of the transthyretin (TTR) gene (V122I), present in 3.43% of African American individuals. Objective:To examine whether serum retinol-binding protein 4 (RBP4), an endogenous TTR ligand, could be used as a diagnostic test for ATTR V122I amyloidosis. Design, Setting, and Participants:In this combined prospective and retrospective cohort study performed at a tertiary care referral center, 50 African American patients 60 years or older with nonamyloid HF and cardiac wall thickening prospectively genotyped from September 1, 2014, through December 31, 2015, and a comparator cohort of 25 patients with biopsy-proven ATTR V122I amyloidosis recruited from September 1, 2009, through November 31, 2014, comprised the development cohort. Twenty-seven African American patients and 9 patients with ATTR V122I amyloidosis comprised the validation cohort. Main Outcomes and Measures:Circulating RBP4, TTR, B-type natriuretic peptide (BNP), and troponin I (TnI) concentrations and electrocardiographic, echocardiographic, and clinical characteristics were assessed in all patients. Receiver operating characteristic (ROC) analysis was performed to identify optimal thresholds for ATTR V122I amyloidosis identification. A clinical prediction rule was developed using penalized logistic regression, evaluated using ROC analysis and validated in an independent cohort of cases and controls. Results:Age, sex, and BNP and TnI concentrations were similar between the 25 patients with ATTR V122I amyloidosis (mean [SD] age, 72.2 [7.4] years; 18 male [72%]) and the 50 controls (mean [SD] age, 69.2 [5.7] years; 31 male [62%]). Serum RBP4 concentration was lower in patients with ATTR V122I amyloidosis compared with nonamyloid controls (31.5 vs 49.4 µg/mL, P?<?.001), and the difference persisted after controlling for potential confounding variables. Left ventricular ejection fraction was lower in patients with ATTR V122I amyloidosis (mean [SD], 40% [14%] vs 57% [14%], P?<?.001), whereas interventricular septal diameter was higher (mean [SD], 16 [3] vs 14 [2] mm, P?<?.001). The ROC analysis identified RBP4 as a sensitive identifier of ATTR V122I amyloidosis (area under the curve [AUC]?=?0.78; 95% CI, 0.67-0.88). A clinical prediction algorithm composed of RBP4, TTR, left ventricular ejection fraction, interventricular septal diameter, mean limb lead QRS voltage, and grade 3 diastolic dysfunction yielded excellent discriminatory capacity for ATTR V122I amyloidosis (AUC?=?0.97; 95% CI, 0.93-1.00), whereas a 4-parameter model, including RBP4 concentration, retained excellent discrimination (AUC?=?0.92; 95% CI, 0.86-0.99). The models maintained excellent discrimination in the validation cohort. Conclusions and Relevance:A prediction model using circulating RBP4 concentration and readily available clinical parameters accurately discriminated ATTR V122I amyloidosis from nonamyloid HF in a case-matched cohort. This clinical algorithm may be useful for identification of ATTR V122I amyloidosis in elderly African American patients with HF.

Dataset Information

Novel Zn2+-binding sites in human transthyretin: implications for amyloidogenesis and retinol-binding protein recognition.

Publications

Novel Zn2+-binding sites in human transthyretin: implications for amyloidogenesis and retinol-binding protein recognition.

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