Reduction of adenosine-5'-phosphosulfate instead of 3'-phosphoadenosine-5'-phosphosulfate in cysteine biosynthesis by Rhizobium meliloti and other members of the family Rhizobiaceae.
We have cloned and sequenced three genes from Rhizobium meliloti (Sinorhizobium meliloti) that are involved in sulfate activation for cysteine biosynthesis. Two of the genes display homology to the Escherichia coli cysDN genes, which code for an ATP sulfurylase (EC 2.7.7.4). The third gene has homology to the E. coli cysH gene, a 3'-phosphoadenosine-5'-phosphosulfate (PAPS) reductase (EC 1.8.99.4), but has greater homology to a set of genes found in Arabidopsis thaliana that encode an adenosine-5'-phosphosulfate (APS) reductase. In order to determine the specificity of the R. meliloti reductase, the R. meliloti cysH homolog was histidine tagged and purified, and its specificity was assayed in vitro. Like the A. thaliana reductases, the histidine-tagged R. meliloti cysH gene product appears to favor APS over PAPS as a substrate, with a Km for APS of 3 to 4 microM but a Km for PAPS of >100 microM. In order to determine whether this preference for APS is unique to R. meliloti among members of the family Rhizobiaceae or is more widespread, cell extracts from R. leguminosarum, Rhizobium sp. strain NGR234, Rhizobium fredii (Sinorhizobium fredii), and Agrobacterium tumefaciens were assayed for APS or PAPS reductase activity. Cell extracts from all four species also preferentially reduce APS over PAPS.
Abola
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AP
Howard Hughes Medical Institute, Department of Biological Sciences, Stanford University, Stanford, California 94305-5020, USA.
Willits
M G
MG
Wang
R C
RC
Long
S R
SR
D000250:Adenosine Phosphosulfate / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D003545:Cysteine / Q000096:biosynthesis*; D004269:DNA, Bacterial; D006639:Histidine; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes; D010084:Oxidation-Reduction; D010724:Phosphoadenosine Phosphosulfate / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012230:Rhizobiaceae / Q000201:enzymology; D017386:Sequence Homology, Amino Acid; D016962:Sinorhizobium meliloti / Q000201:enzymology* / Q000235:genetics; D013447:Sulfites; D015238:Sulfotransferases / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D009097:Multienzyme Complexes; D011993:Recombinant Fusion Proteins; D013447:Sulfites; C075082:nodulation protein Q, Rhizobium; D010724:Phosphoadenosine Phosphosulfate; D000250:Adenosine Phosphosulfate; D006639:Histidine; C022321:PAPS sulfotransferase; D017853:Phosphotransferases (Alcohol Group Acceptor); C019617:adenylylsulfate kinase; D015238:Sulfotransferases; D003545:Cysteine