Cryo-EM structure of native human thyroglobulin

Ricardo Adaixo,Henning Stahlberg,Ricardo D Righetto,Eva M Steiner,Nicholas M I Taylor,Alexander Schmidt

doi:10.1038/s41467-021-27693-8

Abstract

The thyroglobulin (TG) protein is essential to thyroid hormone synthesis, plays a vital role in the regulation of metabolism, development and growth and serves as intraglandular iodine storage. Its architecture is conserved among vertebrates. Synthesis of triiodothyronine (T3) and thyroxine (T4) hormones depends on the conformation, iodination and post-translational modification of TG. Although structural information is available on recombinant and deglycosylated endogenous human thyroglobulin (hTG) from patients with goiters, the structure of native, fully glycosylated hTG remained unknown. Here, we present the cryo-electron microscopy structure of native and fully glycosylated hTG from healthy thyroid glands to 3.2 Å resolution. The structure provides detailed information on hormonogenic and glycosylation sites. We employ liquid chromatography–mass spectrometry (LC-MS) to validate these findings as well as other post-translational modifications and proteolytic cleavage sites. Our results offer insights into thyroid hormonogenesis of native hTG and provide a fundamental understanding of clinically relevant mutations.

Highlights

The thyroglobulin (TG) protein is essential to thyroid hormone synthesis, plays a vital role in the regulation of metabolism, development and growth and serves as intraglandular iodine storage
We identified 57 disulfide bridges (DSB) over the overall human thyroglobulin (hTG) structure (Supplementary Fig. 1 and Supplementary Table 3) stabilizing the protein and three types of cysteine-rich internal homology repeats in hTG: type 1, type 2 and type 3 (Supplementary Table 2)
Both models are overall in close agreement, certain portions of the structure, in particular at the “wing” and the “foot” are better resolved and in some regions deviate from the previous assignment by Coscia et al We describe posttranslational modifications of hTG (Table 1)

Summary

Introduction

The thyroglobulin (TG) protein is essential to thyroid hormone synthesis, plays a vital role in the regulation of metabolism, development and growth and serves as intraglandular iodine storage. Synthesis of triiodothyronine (T3) and thyroxine (T4) hormones depends on the conformation, iodination and post-translational modification of TG. We present the cryo-electron microscopy structure of native and fully glycosylated hTG from healthy thyroid glands to 3.2 Å resolution. We present the structure of endogenous, fully glycosylated hTG determined by cryogenic transmission electron microscopy (cryo-EM). We provide a comprehensive structural description of the endogenous hTG dimer and demonstrate the functional importance of the natural post-translational modification and iodination sites by presenting an atomic model of the nearly complete protein. The native hTG sample is heterogeneous both in composition and conformational states, which likely represents the in vivo requirements for hTG function

Methods

Results

Conclusion