Structure-function analysis of yeast Grx5 monothiol glutaredoxin defines essential amino acids for the function of the protein
Data de publicació2002
Rodríguez Manzanaque, Maria Teresa
MetadadesMostra el registre d'unitat complet
Grx5 defines a family of yeast monothiol glutaredox- ins that also includes Grx3 and Grx4. All three proteins display significant sequence homology with proteins found from bacteria to humans. Grx5 is involved in iron/ sulfur cluster assembly at the mitochondria, but the function of Grx3 and Grx4 is unknown. Three-dimen- sional modeling based on known dithiol glutaredoxin structures predicted a thioredoxin fold structure for Grx5. Positionally conserved amino acids in this glu- taredoxin family were replaced in Grx5, and the effect on the biological function of the protein has been tested. For all changes studied, there was a correlation between the effects on several different phenotypes: sensitivity to oxidants, constitutive protein oxidation, ability for respiratory growth, auxotrophy for a number of amino acids, and iron accumulation. Cys60 and Gly61 are essen- tial for Grx5 function, whereas other single or double substitutions in the same region had no phenotypic ef- fects. Gly115 and Gly116 could be important for the for- mation of a glutathione cleft on the Grx5 surface, in contrast to adjacent Cys117. Substitution of Phe50 alters the ␤-sheet in the thioredoxin fold structure and inhib- its Grx5 function. None of the substitutions tested affect the structure at a significant enough level to reduce protein stability.