Sulfation, the up-and-coming post-translational modification: its role and mechanism in protein-protein interaction

Abstract

Tyrosine sulfation is a post-translational modification entailing covalent attachment of sulfate to tyrosine residues. It takes place in the trans-Golgi, is necessary for the bioactivity of some proteins, and improves their ability to interact with other proteins. In the present work, we show that a protein containing a sulfated tyrosine with a delocalized negative charge forms a salt bridge with another protein if it has two or more adjacent arginine residues containing positive delocalized charges. These noncovalent complexes are so stable that, when submitted to collision induced dissociation, the peptides forming the complex dissociate. Just one covalent bond fragments, the covalent bond between the tyrosine oxygen and the SO3 sulfur, and is represented by the appearance of a new peak (basic peptide + SO3), suggesting that in some instances covalent bonds will break down before the noncovalent bonds between the arginine guanidinium and SO3 dissociate. The data implies that the dissociation pathway is preferred; however, fragmentation between tyrosine and the sulfate residue is a major pathway.

Figure 1

(A) Spectrum showing a noncovalent complex of sulfated leuenkephalin and Dyn 1–8. (B) Spectrum showing a noncovalent complex of sulfated leuenkephalin and Dyn 1–17.

Figure 2

(A) Spectrum showing a noncovalent complex of sulfated cholecystokinin fragment and Dyn 1–8. (B) Spectrum showing a noncovalent complex of sulfated cholecystokinin fragment and Dyn 1–17.

Figure 3

(A) Spectrum showing a noncovalent complex of sulfated hirudin fragment and Dyn 1–8. (B) Spectrum showing a noncovalent complex of sulfated hirudin fragment and Dyn 1–17.

Figure 4

MS/MS spectrum of the complexes of Dyn 1–8 and (A) sulfated leuenkephalin, (B) sulfated cholecystokinin fragment, (C) sulfated hirudin fragment. All spectra show that CID lead to a dissociation of the complex and the appearance of an additional peak at amu 1061 (Dyn 1–8 + SO₃), demonstrating that the noncovalent bond between the arginine guanidinium and the sulfate could be more stable than the covalent bond between the tyrosine oxygen and the sulfate S.

Figure 5

MS/MS spectrum of the complexes of Dyn 1–17 and (A) sulfated leuenkephalin, (B) sulfated cholecystokinin fragment, (C) sulfated hirudin fragment. All spectra show that CID leads to a dissociation of the complex and the appearance of an additional peak at amu 2227 (Dyn 1–17 + SO₃), demonstrating that the noncovalent bond between the arginine guanidinium and the sulfate could be more stable than the covalent bond between the tyrosine oxygen and the sulfate S.