Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0348321 (Haemophilus)
 15,372 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Escherichia coli selenophosphate synthetase (SPS, the selD gene product) catalyzes the production of monoselenophosphate, the selenium donor compound required for synthesis of selenocysteine (Sec) and seleno-tRNAs. We report the molecular cloning of human and mouse homologs of the selD gene, designated Sps2, which contains an in-frame TGA codon at a site corresponding to the enzyme's putative active site. These sequences allow the identification of selD gene homologs in the genomes of the bacterium Haemophilus influenzae and the archaeon Methanococcus jannaschii, which had been previously misinterpreted due to their in-frame TGA codon. Sps2 mRNA levels are elevated in organs previously implicated in the synthesis of selenoproteins and in active sites of blood cell development. In addition, we show that Sps2 mRNA is up-regulated upon activation of T lymphocytes and have mapped the Sps2 gene to mouse chromosome 7. Using the mouse gene isolated from the hematopoietic cell line FDCPmixA4, we devised a construct for protein expression that results in the insertion of a FLAG tag sequence at the N terminus of the SPS2 protein. This strategy allowed us to document the readthrough of the in-frame TGA codon and the incorporation of 75Se into SPS2. These results suggest the existence of an autoregulatory mechanism involving the incorporation of Sec into SPS2 that might be relevant to blood cell biology. This mechanism is likely to have been present in ancient life forms and conserved in a variety of living organisms from all domains of life.
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PMID:Identification of a novel selD homolog from eukaryotes, bacteria, and archaea: is there an autoregulatory mechanism in selenocysteine metabolism? 898 68

The selD gene from Haemophilus influenzae has been overexpressed in Escherichia coli. The expressed protein was purified to homogeneity in a four-step procedure and then carboxymethylated by reaction with chloroacetate. N-terminal sequencing by Edman degradation identified residue 16 as carboxymethyl selenocysteine, which corresponded to the essential cysteine residue in the glycine-rich sequence of the E. coli selenophosphate synthetase. It would be expected that an ionized selenol of a selenocysteine in place of a catalytically essential cysteine residue would result in an enzyme with increased catalytic activity. To test this hypothesis we kinetically characterized the selenocysteine containing selenophosphate synthetase from H. influenzae and compared its catalytic activity to that of the cysteine containing selenophosphate synthetase from E. coli. Our characterization revealed the Km values for the two substrates, selenide and ATP, were similar for both enzymes. However, the selenocysteine-containing enzyme did not exhibit the expected higher catalytic activity. Based on these results we suggest a role of selenocysteine in H. influenzae that is not catalytic.
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PMID:Catalytic properties of selenophosphate synthetases: comparison of the selenocysteine-containing enzyme from Haemophilus influenzae with the corresponding cysteine-containing enzyme from Escherichia coli. 987 69