Cloning and characterization of the NAD-linked glycerol-3-phosphate dehydrogenases of Trypanosoma brucei brucei and Leishmania mexicana mexicana and expression of the trypanosome enzyme in Escherichia coli

Mol Biochem Parasitol. 1996 Feb-Mar;76(1-2):159-73. doi: 10.1016/0166-6851(95)02556-1.


A polyclonal antiserum raised against the purified glycosomal glycerol-3-phosphate dehydrogenase of Trypanosoma brucei brucei has been used to identify the corresponding cDNA clone in a T.b. brucei expression library. This cDNA was subsequently used to obtain genomic clones containing glycerol-3-phosphate dehydrogenase genes. Two tandemly arranged genes were detected in these clones. Characterization of one of the genes showed that it codes for a polypeptide of 353 amino acids, with a molecular mass of 37,651 Da and a calculated net charge of +8. Using the T.b. brucei gene as a probe, a corresponding glycerol-3-phosphate dehydrogenase gene was also identified in a genomic library of Leishmania mexicana mexicana. The L.m. mexicana gene codes for a polypeptide of 365 amino acids, with a molecular mass of 39,140 Da and a calculated net charge of +8. The amino-acid sequences of both polypeptides are 63% identical and carry a type-1 peroxisomal targeting signal (PTS1) SKM and -SKL at their respective C-termini. Moreover, the L.m. mexicana polypeptide also carries a short N-terminal extension reminiscent of a mitochondrial transit sequence. Subcellular localisation analysis showed that in L.m. mexicana the glycerol-3-phosphate dehydrogenase activity co-fractionated both with mitochondria and with glycosomes. This is not the case in T. brucei, where the enzyme is predominantly glycosomal. The two trypanosomatid sequences resemble their prokaryotic homologues (32-36%) more than their eukaryotic counterparts (25-31%) and carry typical prokaryotic signatures. The possible reason for this prokaryotic nature of a trypanosomatid glycerol-3-phosphate dehydrogenase is discussed.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Chromosome Mapping
  • Cloning, Molecular
  • Gene Expression
  • Glycerol-3-Phosphate Dehydrogenase (NAD+)
  • Glycerolphosphate Dehydrogenase / chemistry*
  • Glycerolphosphate Dehydrogenase / genetics
  • Leishmania mexicana / enzymology*
  • Leishmania mexicana / genetics
  • Molecular Sequence Data
  • NAD / metabolism
  • Phylogeny
  • Sequence Homology
  • Trypanosoma brucei brucei / enzymology*
  • Trypanosoma brucei brucei / genetics


  • NAD
  • Glycerolphosphate Dehydrogenase
  • Glycerol-3-Phosphate Dehydrogenase (NAD+)

Associated data

  • GENBANK/X89738
  • GENBANK/X89739
  • GENBANK/X89770