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Introduction

Human thyroglobulin (TG) is a precursor of two thyroid hormones (TH): tetraiodothyronine or thyroxine (T4) and triiodothyronine (T3), essential for growth, development and the control of metabolism. Its structure is essential for the diagnosis, treatment and monitoring of thyroid-related diseases.

PDBID: 6scj.

See the complete sequence.

Composition and 3D structure ^[1]

Global structure

Human thyroglobulin is a dimeric glycoprotein (see a ) consisting of 2 times 2749 amino acid residues with a molecular weight of 660 kDa. The average size of the dimer is 120x235 Å.

Each of its monomers comprises 5 distinct regions on which approximately 66 (chain A in blue, chain B in green, chain C in dark red, chain D in light red) are distributed. These are the regions :

[>NTD ], [>CORE ], [>FLAP ], [>ARM ] and [>CTD ].

The of TG (about 520 amino acids) shows homology with the Acetylcholinesterase and other esterases.

In addition, has about 120 cysteine residues allowing the formation of about 60 per [>Disulfide bridge]. It confers great stability and solubility.

These enzymes belong to the class of α-β hydrolase fold superfamily, characterized by α–helices and β-strands that roughly alternate along the polypeptide chain.^[2]

Structure of hormonogenic sites

Hormonogenic sites are substrates for TH formation. They are formed by 2 or even 3 tyrosines at less than 15 Å from each other, and exposed to the solvent. Of these tyrosines, 1 or 2 are acceptors and 1 is a donor.

There are 4 hormonogenic sites for  :

- an , comprising two fixed donor tyrosines Y234 and Y149 and one flexible acceptor tyrosine: Y24 [>Site A].

- a , including one Y2573 donor and one Y2540 acceptor [>Site B].

- a C site, comprising both donor and acceptor tyrosine Y2766 (not resolved in the map).

- a , comprising a Y108 donor and a Y1310 acceptor [>Site D].

An acidic Asp (aspartic acid) or Glu (glutamic acid) residue always precedes the acceptor and a lysine is always close to the donor.

Except for site C, donors are at the fixed regions of the dimer while acceptors are at the flexible regions.

Synthesis^[3][4][5] and acquisition of its structure

Role and composition of the thyroid gland

The thyroid gland is an endocrine organ located in the neck that secretes thyroid hormones into the bloodstream. Among them, T4 and T3 are involved in the growth, development and regulation of the metabolism of vertebrates.

The thyroid gland is made up of thyroid vesicles, which are made up of thyroid follicular cells that are arranged around a lumen containing a viscous substance called colloid. The diet is a source of iodide I- ions which, once in the blood, are picked up by the thyroid cells and then partly discharged into the colloid.

Expression of the TG gene

In humans, TG is coded by a large gene roughly 300 kb long, located on chromosome 8. The number of exons has been estimated to be around 48, each of which is separated by introns varying in size up to 64 kb. TG gene expression is controlled positively by thyro-tropin (TSH) through the modulation of the intra-cellular levels of cyclic adenosine monophosphate (cAMP) via its receptor (TSHr) located at the basal membrane of the cell.^[6]

Post-translational modifications

TG also undergoes N-glycosylations in the endoplasmic reticulum at , so that 10% of its molecular weight is carbohydrate. TG also undergoes maturation steps in this organelle, where it acquires dilsufide-bonds. These modifications enhance its stability and solubility. Indeed, the two monomers are linked not by covalent interactions but via numerous interactions allowed by these N-glycosylations.

Folding

Once synthesized, the acquisition of the 3D structure of the [>TG ] is enabled by the ER chaperone proteins of the thyroid follicular cells via a slow process ^[7].

Functions

TH synthesis

Mechanism

Once their 3D structure is acquired, TGs are exported into the colloid by exocytosis thanks to their signal peptide which will be cleaved. This extracellular storage increases the amount of TG stored in the body.

In the colloid, about 30 tyrosines out of the 66 tyrosines, each consisting of a phenol group, are iodized. The quantity of iodinated tyrosine depends however on the iodide concentration of the colloid. Indeed, one or two iodide ions can be covalently bound to the colloid and thus give a di- (DIT) or mono-iodinated (MIT) phenol group. The iodination of the phenol groups is carried out by two membrane enzymes of the follicular cells: the double oxidase (DUOX) synthesizes the hydrogen peroxide H2O2 necessary for thyroid peroxidase (TPO). The synthesis is completed by TG proteolysis.

Due to the spatial conformation of , there is a transfer of di- or mono-iodinated aromatic ring from a donor tyrosine to a close acceptor diiodotyrosine for the 14 tyrosines of the hormonogenic sites. Acceptor iodinated tyrosines are DITs because they are deprotonated due to their 6.5 acid pka facilitating the acceptance reaction leading to the formation of quinol-ether bonds, whereas donor iodinated tyrosines are MITs with a pKa of 8.5^[8].

At the end of the coupling, the donor tyrosines are left with a dehydroalanine.

Once iodization and coupling have been performed, endocytosis of the colloid to the lysosome occurs. The is proteolyzed by cathepsin proteases^[9] and 7 TH are thus released from 14 mono- or di-iodinated tyrosines.

The hormone synthesis function of TG is thus particularly linked to its structure. Moreover, research shows that denaturation or a simple modification of its conformation prevents the formation of TH ^[10][11].

Control

The synthesis of TH from TG is stimulated by thyroid stimulating hormone (TSH) secreted by the pituitary gland, a gland of the brain. When the TSH receptor is activated, glycosylations leading to the mono iodination of tyrosines promote the synthesis of T3^[12][13]. On the other hand, if the amount of hormones is too high, a negative feedback is exerted on this process while a small amount of these hormones exerts a positive feedback.

Iodine tank

The complex and particularly stable structure of gives it iodide reservoir properties. Indeed, all iodinated but non-hormonoid tyrosines are useful for iodine storage in the thyroid gland.

Interest in the medical field^[14][15][2]

Modification of the TG quantity related to the disease

A healthy subject has between 5 and 25 µg of TG per liter of blood.^[16] In case of thyroid dysfunction, this level may increase or decrease. For example, it decreases in the case of congenital athyreosis (insufficiency of the thyroid gland) or prior to a miscarriage due to the presence of anti-TG antibodies, but increases in the case of cancer, thyroiditis, inflammation of the thyroid or autoimmune thyroid diseases AITD ^[17](Grave's disease, Hashimoto's thyroiditis).^[18][19]

In addition, a decrease in the size or capacity of the thyroid causes a decrease in the synthesis of TH by the , and thus a drop in the blood level of T4 and T3, which in turn causes heart disease, brain disease and abnormalities in the development of the fetus. ^[20][21][22]

The variation of the quantity of TG can thus be as much a cause as a consequence of disease.^[23]

Use of TG to treat diseases

A classic TSH-stimulated measurement or ultrasensitive measurement allows to control its rate in a more or less sensitive way and therefore to detect a disease like those mentioned above, to ensure the effectiveness of a treatment and the absence of recurrence and to avoid miscarriages.^[24] Since serum levels are correlated with the volume of thyroid tissue, we can also estimate the mass of thyroid tissue to detect hyperthyroidism, a disease related to an enlarged thyroid or, conversely, hypothyroidism.

For example, thyroidectomy and a iodine-131 therapy can be performed to cure thyroid cancer with an 80% chance. Following removal and iodine-131 therapy, thyroglobulin is this time produced by malignant thyrocytes. As a result, its blood level is indistinguishable from that of a healthy person. ^[25][26]

But in case of recurrence and persistence of cancer, its level can increase again. Thyroglobulin therefore always serves as a tumor marker allowing us to estimate the risk of recurrence (>2ng/ml) or persistence (>1ng/ml) or remission.^[27]