Hereditary Lysozyme Amyloidosis Variant p.Leu102Ser Associates with Unique Phenotype

Abstract

Lysozyme amyloidosis (ALys) is a rare form of hereditary amyloidosis that typically manifests with renal impairment, gastrointestinal (GI) symptoms, and sicca syndrome, whereas cardiac involvement is exceedingly rare and neuropathy has not been reported. Here, we describe a 40-year-old man with renal impairment, cardiac and GI symptoms, and peripheral neuropathy. Renal biopsy specimen analysis revealed amyloidosis with extensive involvement of glomeruli, vessels, and medulla. Amyloid was also detected in the GI tract. Echocardiographic and electrocardiographic findings were consistent with cardiac involvement. Proteomic analysis of Congo red-positive renal and GI amyloid deposits detected abundant lysozyme C protein. DNA sequencing of the lysozyme gene in the patient and his mother detected a heterozygous c.305T>C alteration in exon 3, which causes a leucine to serine substitution at codon 102 (Human Genome Variation Society nomenclature: p.Leu102Ser; legacy designation: L84S). We also detected the mutant peptide in the proband's renal and GI amyloid deposits. PolyPhen analysis predicted that the mutation damages the encoded protein. Molecular dynamics simulations suggested that the pathogenesis of ALys p.Leu102Ser is mediated by shifting the position of the central β-hairpin coordinated with an antiparallel motion of the C-terminal helix, which may alter the native-state structural ensemble of the molecule, leading to aggregation-prone intermediates.

Keywords: amyloidosis; hereditary amyloid; lysozyme; malfolding proteins.

Figure 1.

Renal and GI biopsy findings. (A) Abundant glomerular mesangial, glomerular capillary wall, and arterial congophilic amyloid deposits are seen. (B) Renal CR-positive amyloid deposits display apple-green birefringence under polarized light. (C) Extensive congophilic amyloid deposits are seen in the medulla involving collecting ducts and vasa recta basement membranes. (D) On electron microscopy, amyloid deposits are composed of randomly oriented straight fibrils. (E) The intestinal mucosal amyloid deposits stain strongly for lysozyme using immunohistochemistry. Paneth cells at the bases of the glands serve as an internal positive control for lysozyme. Proteomic data confirming the diagnosis of ALys with the p.Leu102Ser abnormality from this specimen are presented in Supplemental Figure 1. Magnification, ×100 in A–C and E; ×50,000 in D.

Figure 2.

Proteomic and genetic detection of amyloidosis derived from lysozyme Leu102Ser variant. (A) Proteins highlighted with yellow stars are the universal amyloid tissue biomarkers. LYZ protein is highlighted with a blue star. Numbers in the boxes represent the total numbers of spectra matched to the protein in a sample. Multiple dissections are presented independently. (B) Portions of the LYZ sequence detected in the sample are highlighted with bold black letters on yellow background. (C) Tandem mass spectrometry spectrum of the mutant peptide detected in the CR-positive renal amyloid deposits of the patient. An arrow marks the p.Leu102Ser amino acid sequence change. (D) LYZ gene mutation in the patient. An arrow marks the detected c.305T→C (p.Leu102Ser) mutation.

Figure 3.

Family tree of the affected kindred. The proband is indicated in red. Dead individuals are indicated by a diagonal line through the symbol. Miscarriages are indicated with diamonds.

Figure 4.

MD of ALys p.Leu102Ser. (A) Previously reported ALys coding variants are marked by black spheres. Newly observed p.Leu102Ser variant site is indicated in orange text. Previously reported variants are either within the same sequence-based region of LYZ or closely interact with them in three dimensions. p.Leu102Ser is distant in sequence but close in space to previously reported amyloidosis variants. (B) The root mean squared deviation (RMSD) across replicates and for each variant is shown by a smoothed density plot, where the width of each shape is proportional to the number of observations. The WT is summarized in blue, the novel L102S is in red, and three previously identified amyloidosis variants are in shades of orange. (C) PCs were computed across all 15 simulations. The first PC is visualized on the LYZ structure. The final WT conformation is shown in cartoon representation. Amino acid 102 is marked by a red sphere, whereas 72, 74, and 75 are marked by orange spheres. Two terminal amino acids exhibited large motions and are not shown. The first PC indicated anticorrelated motion between the β-hairpin and C-terminal α-helices. Both known and novel ALys variants destabilize LYZ protein by introducing flexibility in the C terminus of the protein.