Abstract
BackgroundGrx6 is a yeast Golgi/endoplasmic reticulum protein involved in iron-sulfur binding that belongs to monothiol glutaredoxin-protein family. Grx6 has been biochemically characterized previously. Grx6 contains a conserved cysteine residue (Cys-136). Depending on the active-site sequences, Grxs can be classified to classic dithiol Grxs with a CXXC motif known as classes II and monothiol Grxs with a CXXS motif known as classes I, and Grx6 belongs to the class I with a CSYS motif.ResultsOur results showed how the loop between the N-terminal and C-terminal can affect the stability. When Grx6 was incubated with FeSO4·7H2O and (NH4)2Fe(SO4)2·6H2O, a disulfide bond was formed between the cysteine 136 and glutathione, and the concentration of dimer and tetramer was increased. The results presented various levels of stability of Grx6 with mutant and deleted amino acids. We also highlighted the difference between the monomer and dimer forms of the Grx6, in addition to comparison of the Fe-S cluster positions among holo forms of poplar Grx-C1, human Grx2 and Saccharomyces cerevisiae Grx6.ConclusionsIn this paper, we used a combination of spectroscopic and proteomic techniques to analyse the sequence and to determine the affected mutations and deletions in the stability of Grx6. Our results have increased the knowledge about the differences between monomer and dimer structures in cellular processes and proteins whose roles and functions depend on YCA1 in yeast.
Highlights
Grx6 is a yeast Golgi/endoplasmic reticulum protein involved in iron-sulfur binding that belongs to monothiol glutaredoxin-protein family
Gel filtration Glutaredoxin proteins (Grx) have a common active site motif CxxC/S located at a surface loop that is accessible to GSH
We propose some mutations on the loop between the N and C domains significantly interact with the Fe-S and the GSH dependent oxidoreductase activity
Summary
Grx is a yeast Golgi/endoplasmic reticulum protein involved in iron-sulfur binding that belongs to monothiol glutaredoxin-protein family. Grx contains a conserved cysteine residue (Cys-136). Depending on the active-site sequences, Grxs can be classified to classic dithiol Grxs with a CXXC motif known as classes II and monothiol Grxs with a CXXS motif known as classes I, and Grx belongs to the class I with a CSYS motif. Glutaredoxin proteins (Grx) have thiol reductase activity. Glutaredoxins are necessary to reduce glutathione (GSH) as the electron donor [1]. The assumptive glutaredoxin sequences consist of an active site that contains one or two conserved cysteines together with two non-conserved residues among them. Previous studies have shown that two conserved cysteines are necessary for reducing protein disulfides. Only the cysteine locus at the N-terminal is essential for the reduction of incorporated disulfides with glutathione.
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