cDNA cloning, purification, and characterization of mouse liver selenocysteine lyase. Candidate for selenium delivery protein in selenoprotein synthesis

J Biol Chem. 2000 Mar 3;275(9):6195-200. doi: 10.1074/jbc.275.9.6195.

Abstract

Selenocysteine lyase (SCL) (EC 4.4.1.16) is a pyridoxal 5'-phosphate-dependent enzyme that specifically catalyzes the decomposition of L-selenocysteine to L-alanine and elemental selenium. The enzyme was proposed to function as a selenium delivery protein to selenophosphate synthetase in selenoprotein biosynthesis (Lacourciere, G. M., and Stadtman, T. C. (1998) J. Biol. Chem. 273, 30921-30926). We purified SCL from pig liver and determined its partial amino acid sequences. Mouse cDNA clones encoding peptides resembling pig SCL were found in the expressed sequence tag data base, and their sequences were used as probes to isolate full-length mouse liver cDNA. The cDNA for mouse SCL (mSCL) was determined to be 2,172 base pairs in length, containing an open reading frame encoding a polypeptide chain of 432 amino acid residues (M(r) 47, 201). We also determined the sequence of the N-terminal region of putative human SCL. These enzymes were shown to be distantly related in primary structure to NifS, which catalyzes the desulfurization of L-cysteine to provide sulfur for iron-sulfur clusters. The recombinant mSCL overproduced in Escherichia coli was a homodimer with the subunit M(r) of 47,000. The enzyme was pyridoxal phosphate-dependent and highly specific to L-selenocysteine (the k(cat)/K(m) value for L-selenocysteine was about 4,200 times higher than that for L-cysteine). Reverse transcriptase-polymerase chain reaction and Western blot analyses revealed that mSCL is cytosolic and predominantly exists in the liver, kidney, and testis, where mouse selenophosphate synthetase is also abundant, supporting the view that mSCL functions in cooperation with selenophosphate synthetase in selenoprotein synthesis. This is the first report of the primary structure of mammalian SCL.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Cytosol / enzymology
  • Dimerization
  • Escherichia coli
  • Kinetics
  • Liver / enzymology*
  • Lyases / chemistry
  • Lyases / genetics*
  • Lyases / isolation & purification
  • Mice
  • Molecular Sequence Data
  • Phylogeny
  • Protein Biosynthesis*
  • Proteins*
  • Pyridoxal Phosphate / pharmacology
  • RNA, Messenger
  • Recombinant Proteins / chemistry
  • Selenium / metabolism*
  • Selenoproteins
  • Sequence Alignment
  • Substrate Specificity

Substances

  • Proteins
  • RNA, Messenger
  • Recombinant Proteins
  • Selenoproteins
  • Pyridoxal Phosphate
  • Lyases
  • selenocysteine lyase
  • Selenium

Associated data

  • GENBANK/AF175407
  • GENBANK/AF175767