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. 1986 Jul;83(13):4650–4654. doi: 10.1073/pnas.83.13.4650

Nucleotide sequence and expression of the selenocysteine-containing polypeptide of formate dehydrogenase (formate-hydrogen-lyase-linked) from Escherichia coli.

F Zinoni, A Birkmann, T C Stadtman, A Böck
PMCID: PMC323799  PMID: 2941757

Abstract

The gene (fdhF) coding for the selenopolypeptide of the benzylviologen-linked formate dehydrogenase of Escherichia coli was cloned and its nucleotide sequence was determined. The fdhF gene contains, within an open reading frame coding for a protein of 715 amino acids (calculated molecular weight, 79,087), an opal (UGA) nonsense codon in amino acid position 140. Existence of this nonsense codon was confirmed by physical recloning and resequencing. Internal and terminal deletion clones and lacZ fusions of different N-terminal parts of fdhF were constructed and analyzed for selenium incorporation. Selenylated truncated polypeptide chains or beta-galactosidase fusion proteins were synthesized when the deletion clones or gene fusions, respectively, contained the fdhF gene fragment coding for the selenopolypeptide sequence from amino acid residue 129 to amino acid residue 268. Translation of the lacZ part of the fusions required the presence of selenium in the medium when the N-terminal fdhF part contained the UGA codon and was independent of the presence of selenium when a more upstream part of fdhF was fused to lacZ. The results are consistent with a co-translational selenocysteine incorporation mechanism.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adman E. T., Sieker L. C., Jensen L. H. Structure of a bacterial ferredoxin. J Biol Chem. 1973 Jun 10;248(11):3987–3996. [PubMed] [Google Scholar]
  2. Barrell B. G., Bankier A. T., Drouin J. A different genetic code in human mitochondria. Nature. 1979 Nov 8;282(5735):189–194. doi: 10.1038/282189a0. [DOI] [PubMed] [Google Scholar]
  3. Casadaban M. J., Chou J., Cohen S. N. In vitro gene fusions that join an enzymatically active beta-galactosidase segment to amino-terminal fragments of exogenous proteins: Escherichia coli plasmid vectors for the detection and cloning of translational initiation signals. J Bacteriol. 1980 Aug;143(2):971–980. doi: 10.1128/jb.143.2.971-980.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Casadaban M. J., Cohen S. N. Lactose genes fused to exogenous promoters in one step using a Mu-lac bacteriophage: in vivo probe for transcriptional control sequences. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4530–4533. doi: 10.1073/pnas.76.9.4530. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chang A. C., Cohen S. N. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol. 1978 Jun;134(3):1141–1156. doi: 10.1128/jb.134.3.1141-1156.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cox J. C., Edwards E. S., DeMoss J. A. Resolution of distinct selenium-containing formate dehydrogenases from Escherichia coli. J Bacteriol. 1981 Mar;145(3):1317–1324. doi: 10.1128/jb.145.3.1317-1324.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Craigen W. J., Cook R. G., Tate W. P., Caskey C. T. Bacterial peptide chain release factors: conserved primary structure and possible frameshift regulation of release factor 2. Proc Natl Acad Sci U S A. 1985 Jun;82(11):3616–3620. doi: 10.1073/pnas.82.11.3616. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gray C. P., Sommer R., Polke C., Beck E., Schaller H. Structure of the orgin of DNA replication of bacteriophage fd. Proc Natl Acad Sci U S A. 1978 Jan;75(1):50–53. doi: 10.1073/pnas.75.1.50. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Gray C. T., Wimpenny J. W., Hughes D. E., Mossman M. R. Regulation of metabolism in facultative bacteria. I. Structural and functional changes in Escherichia coli associated with shifts between the aerobic and anaerobic states. Biochim Biophys Acta. 1966 Mar 28;117(1):22–32. doi: 10.1016/0304-4165(66)90148-6. [DOI] [PubMed] [Google Scholar]
  10. Hawkes W. C., Tappel A. L. In vitro synthesis of glutathione peroxidase from selenite. Translational incorporation of selenocysteine. Biochim Biophys Acta. 1983 Mar 10;739(2):225–234. doi: 10.1016/0167-4781(83)90033-7. [DOI] [PubMed] [Google Scholar]
  11. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  12. Pecher A., Zinoni F., Böck A. The seleno-polypeptide of formic dehydrogenase (formate hydrogen-lyase linked) from Escherichia coli: genetic analysis. Arch Microbiol. 1985 May;141(4):359–363. doi: 10.1007/BF00428850. [DOI] [PubMed] [Google Scholar]
  13. Sancar A., Hack A. M., Rupp W. D. Simple method for identification of plasmid-coded proteins. J Bacteriol. 1979 Jan;137(1):692–693. doi: 10.1128/jb.137.1.692-693.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Stadtman T. C. Selenium-dependent enzymes. Annu Rev Biochem. 1980;49:93–110. doi: 10.1146/annurev.bi.49.070180.000521. [DOI] [PubMed] [Google Scholar]
  15. Wittwer A. J. Specific incorporation of selenium into lysine- and glutamate- accepting tRNAs from Escherichia coli. J Biol Chem. 1983 Jul 25;258(14):8637–8641. [PubMed] [Google Scholar]
  16. Yamao F., Muto A., Kawauchi Y., Iwami M., Iwagami S., Azumi Y., Osawa S. UGA is read as tryptophan in Mycoplasma capricolum. Proc Natl Acad Sci U S A. 1985 Apr;82(8):2306–2309. doi: 10.1073/pnas.82.8.2306. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Young P. A., Kaiser I. I. Aminoacylation of Escherichia coli cysteine tRNA by selenocysteine. Arch Biochem Biophys. 1975 Dec;171(2):483–489. doi: 10.1016/0003-9861(75)90057-0. [DOI] [PubMed] [Google Scholar]

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