Properties and Mechanism Studies of 5′‐Adenylylsulfate Reductases

Abstract

5′‐adenylylsulfate (APS) reductases is essential for cysteine biosynthesis in bacteria and plants catalyze the formation of sulfite from APS using either glutathione or thioredoxin. APS reductases from the bacteria Pseudomonas aeruginosa (PaAPR) and the green alga Enteromorpha intestinalis (EiAPR) and the archaeon Methanocaldococcus janishii were studied. EiAPR has a bipartite primary structure consisting of a domain with homology to PaAPR, and C‐terminal domain with homology to glutaredoxin. Iron and acid‐labile sulfide analysis indicated these enzymes both contain one [4Fe‐4S] cluster. Resonance Raman experiments confirmed the presence of a [4Fe‐4S] cluster. Redox titrations show that the EiAPR contains two disulfide/dithiol pairs with midpoint potential (Em) values of ‐290 mV and ‐140 mV, respectively. PaAPR contains only a single disulfide/dithiol cysteine pair with an Em value identical to that observed for the more negative component found in EiAPR. C‐terminal domain of EiAPR itself shows an Em value (i.e., ‐140 mV), consistent with the more positive EiAPR component, which is closer to that of E.coli glutaredoxin (i.e., ‐198 mV) than it is to E.coli thoredoxins (i.e., ‐270 mV). These results suggest that EiAPR has two different disulfide couples; one is like PaAPR and the other (i.e., the C‐domain site) may function in a manner similar to a glutaredoxin.Supported by a grant from USDA and the Baylor University Undergraduate Research and Scholarly Activities Small Grant Program and the Vice Provost for Research.