Enolase from Trypanosoma brucei, from the Amitochondriate Protist Mastigamoeba balamuthi, and from the Chloroplast and Cytosol of Euglena gracilis: Pieces in the Evolutionary Puzzle of the Eukaryotic Glycolytic Pathway

Abstract

Genomic or cDNA clones for the glycolytic enzyme enolase were isolated from the amitochondriate pelobiont Mastigamoeba balamuthi, from the kinetoplastid Trypanosoma brucei, and from the euglenid Euglena gracilis. Clones for the cytosolic enzyme were found in all three organisms, whereas Euglena was found to also express mRNA for a second isoenzyme that possesses a putative N-terminal plastid-targeting peptide and is probably targeted to the chloroplast. Database searching revealed that Arabidopsis also possesses a second enolase gene that encodes an N-terminal extension and is likely targeted to the chloroplast. A phylogeny of enolase amino acid sequences from 6 archaebacteria, 24 eubacteria, and 32 eukaryotes showed that the Mastigamoeba enolase tended to branch with its homologs from Trypanosoma and from the amitochondriate protist Entamoeba histolytica. The compartment-specific isoenzymes in Euglena arose through a gene duplication independent of that which gave rise to the compartment-specific isoenzymes in Arabidopsis, as evidenced by the finding that the Euglena enolases are more similar to the homolog from the eubacterium Treponema pallidum than they are to homologs from any other organism sampled. In marked contrast to all other glycolytic enzymes studied to date, enolases from all eukaryotes surveyed here (except Euglena) are not markedly more similar to eubacterial than to archaebacterial homologs. An intriguing indel shared by enolase from eukaryotes, from the archaebacterium Methanococcus jannaschii, and from the eubacterium Campylobacter jejuni maps to the surface of the three-dimensional structure of the enzyme and appears to have occurred at the same position in parallel in independent lineages.

On the origins of cells: a hypothesis for the evolutionary transitions from abiotic geochemistry to chemoautotrophic prokaryotes, and from prokaryotes to nucleated cells

“…First, although archaebacteria possess pathways from glucose to pyruvate, they have several alternative pathways that involve different enzymatic steps, hence different enzymes not present in glycolysis (Daniel & Danson 1995;Schö nheit & Schäfer 1995;Kengen et al 1996;Selig et al 1997). These include: (i) the modified Embden-Meyerhof pathway, which employs ADP-instead of ATP-dependent steps and a glyceraldehyde-3-phosphate oxidoreductase in place of the steps catalysed by GAPDH and 3-phosphoglycerate kinase; (ii) the Entner-Doudoroff pathway, which does not require the enzymatic steps catalysed by GPI, PFK, FBA or TPI; and (iii) the non-phosphorylated Entner-Doudoroff pathway, which shares only two enzymatic steps in common with glycolysis (Selig et al 1997), those catalysed by enolase (Hannaert et al 2000) and pyruvate kinase (Schramm et al 2000). Second, several archaebacteria have complete glycolytic pathways, but for many steps they use totally different enzymes that are unrelated to the familiar homologues found in eubacteria (and eukaryotes) (Schö nheit & Schäfer 1995;Selig et al 1997).…”

“…This approach to the holochirality problem implicates the peptidyl transferase reaction of the ribosome as the pacemaker for amino-acid stereochemistry and a simpler precursor of enolase as the pacemaker of sugar stereochemistry. It is probably just curious coincidence that enolase is the only enzyme of carbon metabolism, other than adenylate kinase, that balances AMP, ADP and ATP levels, that is encoded in the highly conserved ribosomal protein superoperon in archaebacteria (Hannaert et al 2000), which might also carry a faint trace of chiral origins. At any rate, our suggestion for the origin of holochirality starts with the peptidyl transferase reaction and is thus largely congruent with Woese's (2002, p. 8745) assessment 'The evolution of modern cells, then, had to begin with the onset of translation'.…”

“…At the same time, similar studies were being conducted on the glycolytic enzymes from amitochondriate protists, as reviewed by Mü ller (1998). For every glycolytic enzyme (except enolase) (Hannaert et al 2000) the same observation was scored: the eukaryotic enzymes are more similar to their eubacterial homologues than they are to their archaebacterial homologues, as shown once more in figure 1 for the glycolytic pathway, and in many cases the archaebacteria do not even possess the homologous enzyme.…”

Euglena gracilis Rhodoquinone:Ubiquinone Ratio and Mitochondrial Proteome Differ under Aerobic and Anaerobic Conditions

“…Identification of PDH Subunits E1␣, E2, and E3-Standard molecular methods, nucleic acid isolation, cDNA synthesis, and cloning in ZAPII were performed as described previously (36,37). Hybridization probes for PDH subunits from Euglena were obtained by comparisons of in-house Euglena EST data with annotated sequences in the National Center for Biotechnology Information data base using BLAST.…”

Phylogeny of 33 ribosomal and six other proteins encoded in an ancient gene cluster that is conserved across prokaryotic genomes: influence of excluding poorly alignable sites from analysis.

“…However, a few proteins belonging to other functional classes, notably adenylate kinase (Sa! nchez & Mu $ ller, 1998) and enolase (Hannaert et al, 2000) involved in energy metabolism, are also found in the cluster in some genomes (Wa$ chtersha$ user, 1998). This cluster has proven useful from the standpoint of plastid phylogeny by virtue of its tendency to preserve gene order, revealing evolutionary relationships among plastids (Stoebe & Kowallik, 1999 (Wa$ chtersha$ user, 1998).…”