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Abstract

The effect of in vitro ultraviolet irradiation at 2537 Å on the structure of highly purified human thyroglobulin has been investigated by following the ultracentrifugal pattern and the appearance of free sulfhydryl groups in the irradiated protein.

Thyroglobulin (19 S) dissociates into two 12 S subunits when it is irradiated with doses of a few hundred Einsteins per mole. The quantum yields of dissociation vary between 0.5 × 10⁻³ and 2 × 10⁻³, depending on the experimental conditions. At higher ultraviolet doses, the protein is partially aggregated and denatured.

The extent of dissociation is largely dependent on the ionic strength and the pH of the medium. Doses up to 1,500 Einsteins per mole cause only a partial (30%) splitting of the 19 S molecules, at neutral pH in 0.1 M KCl, whereas at pH 10 and low salt (Γ/2=0.0042), the same dose produces almost complete dissociation into 12 S subunits.

Ultraviolet light also causes the rupture of some of the disulfide bonds of native thyroglobulin. On irradiation with several hundred Einsteins per mole protein, as many as 21 –SH groups per molecule can be titrated. The quantum yields for –SH production and destruction of cystine residues are both 0.05.

No direct correlation has been found between the extent of dissociation of thyroglobulin and the number of –S–S– bonds cleaved, nor between the reassociation of the 12 S subunits and the reoxidation of –SH groups. It is concluded that two 12 S units present in human thyroglobulin are not linked by interchain disulfide bridges; the disulfides cleaved by ultraviolet irradiation, however, may play an important role in maintaining the subunit organization of the protein molecule.