Glycogen Phosphorylase Sequences from the Amitochondriate Protists, Trichomonas vaginalis, Mastigamoeba balamuthi, Entamoeba histolytica and Giardia intestinalis¹

Abstract

Glycogen phosphorylase genes or messages from four amitochondriate eukaryotes, Trichomonas vaginalis, Mastigamoeba balamuthi, Entamoeba histolytica (two genes) and Giardia intestinalis, have been isolated and sequenced. The sequences of the amitochondriate protist enzymes appear to share a most recent common ancestor. The clade containing these sequences is closest to that of another protist, the slime mold (Dictyostelium discoideum), and is more closely related to fungal and plant phosphorylases than to mammalian and eubacterial homologs. Structure-based amino acid alignment shows conservation of the residues and domains involved in catalysis and allosteric regulation by glucose 6-phosphate but high divergence at domains involved in phosphorylation-dependent regulation and AMP binding in fungi and animals. Protist phosphorylases, as their prokaryotic and plant counterparts, are probably not regulated by phosphorylation.

Glycogen Accumulation and Degradation by the Trichomonads Trichomonas vaginalis and Trichomonas tenax

“…As we had obtained evidence that the glycogen phosphorylases from T. vaginalis and T. tenax were unlikely to require phosphorylation for activation, we reasoned that it might be possible to produce active, recombinant phosphorylase in E. coli . Therefore, we designed PCR primers based on the sequence, now designated TVAG_348330, reported by Wu and Müller () and amplified a product of approximately 2.8 kb from T. vaginalis cDNA. We sequenced this PCR product and found it to be identical to TVAG_348330.…”

“…Wu and Müller () reported cloning a putative glycogen phosphorylase from T. vaginalis . The completion of the T. vaginalis genome sequence revealed that the organism contained a second open reading frame that likely encoded glycogen phosphorylase (Carlton et al.…”

“…Glycogen phosphorylase liberates glucose residues in the form of glucose-1-phosphate sequentially from the nonreducing ends of the a-1,4-linked glucose chains (Roach 2002;Wilson et al 2010), and a separate activity, known as debranching enzyme, is required to eliminate the a-1,6-linked branches. Wu and Mu¨ller (2003) reported cloning a putative glycogen phosphorylase from T. vaginalis. The completion of the T. vaginalis genome sequence revealed that the organism contained a second open reading frame that likely encoded glycogen phosphorylase (Carlton et al 2007).…”

Biochemistry and Evolution of Anaerobic Energy Metabolism in Eukaryotes

“…Abbreviations: Fd, ferredoxin; [1], pyruvate:ferredoxin oxidoreductase; [5], malic enzyme; [9], pyruvate kinase; [11], malate dehydrogenase; [18], alanine aminotransferase; [25], alcohol dehydrogenase E; [28], phosphoenolpyruvate carboxytransferase (PP i dependent); [29], pyruvate:orthophosphate dikinase; [30] Because of the potential of the mitosome to harbor drug targets, protein import into Entamoeba mitosomes has been intensely studied as of late (118). A free-living relative of Entamoeba, Mastigamoeba balmuthii, has several enzymes of core energy metabolism in common with Entamoeba (183,564) and also possesses mitochondria that import Cpn60 (159).…”

Glycogen as a carbohydrate energy reserve in trophozoites of Giardia lamblia

“…Some structures similar to glycogen particles have already been observed in the cytoplasm of trophozoites using electron microscopy (Sogayar and Gregorio 1991;Lanfredi-Rangel et al 1999). Nucleotide fragments of the enzyme glycogen phosophorylase, involved in the degradation of glycogen, were isolated and sequenced from trophozoites (Wu and Muller 2003). In the light of this observation, we sought to purify the high molecular weight carbohydrate present in G. lamblia trophozoites, determining its monosaccharide constituents, concentration relative to total carbohydrate and role as energetic reserve.…”