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At least 145 records · Page 8Linked to original sources
PubMed1999

Characterization of the gene encoding serine acetyltransferase, a regulated enzyme of cysteine biosynthesis from the protist parasites Entamoeba histolytica and Entamoeba dispar. Regulation and possible function of the cysteine biosynthetic pathway in Entamoeba.

The enteric protist parasites Entamoeba histolytica and Entamoeba dispar possess a cysteine biosynthetic pathway, unlike their mammalian host, and are capable of de novo production of L-cysteine. We cloned and characterized cDNAs that encode the regulated enzyme serine acetyltransferase (SAT) in this pathway from these amoebae by genetic complementation of a cysteine-auxotrophic Escherichia coli strain with the amoebic cDNA libraries. The deduced amino acid sequences of the amoebic SATs exhibited, within the most conserved region, 36-52% identities with the bacterial and plant SATs. The amoebic SATs contain a unique insertion of eight amino acids, also found in the corresponding region of a plasmid-encoded SAT from Synechococcus sp., which showed the highest overall identities to the amoebic SATs. Phylogenetic reconstruction also revealed a close kinship of the amoebic SATs with cyanobacterial SATs. Biochemical characterization of the recombinant E. histolytica SAT revealed several enzymatic features that distinguished the amoebic enzyme from the bacterial and plant enzymes: 1) inhibition by L-cysteine in a competitive manner with L-serine; 2) inhibition by L-cystine; and 3) no association with cysteine synthase. Genetically engineered amoeba strains that overproduced cysteine synthase and SAT were created. The cysteine synthase-overproducing amoebae had a higher level of cysteine synthase activity and total thiol content and revealed increased resistance to hydrogen peroxide. These results indicate that the cysteine biosynthetic pathway plays an important role in antioxidative defense of these enteric parasites.

Acetyltransferases↗
PubMed2003

Identification and characterization of two isoenzymes of methionine gamma-lyase from Entamoeba histolytica: a key enzyme of sulfur-amino acid degradation in an anaerobic parasitic protist that lacks forward and reverse trans-sulfuration pathways.

To better understand the metabolism of sulfur-containing amino acids, which likely plays a key role in a variety of cell functions, in Entamoeba histolytica, we searched the genome data base for genes encoding putative orthologs of enzymes known to be involved in the metabolism. The search revealed that E. histolytica possesses only incomplete cysteine-methionine conversion pathways in both directions. Instead, this parasite possesses genes encoding two isoenzymes of methionine gamma-lyase (EC 4.4.1.11, EhMGL1/2), which has been implicated in the degradation of sulfur-containing amino acids. The two amebic MGL isoenzymes, showing 69% identity to each other, encode 389- and 392-amino acid polypeptides with predicted molecular masses of 42.3 and 42.7 kDa and pIs of 6.01 and 6.63, respectively. Amino acid comparison and phylogenetic analysis suggested that these amebic MGLs are likely to have been horizontally transferred from the Archaea, whereas an MGL from another anaerobic protist Trichomonas vaginalis has MGL isotypes that share a common ancestor with bacteria. Enzymological and immunoblot analyses of the partially purified native amebic MGL confirmed that both of the MGL isotypes are expressed in a comparable amount predominantly in the cytosol and form a homotetramer. Recombinant EhMGL1 and 2 proteins catalyzed degradation of L-methionine, DL-homocysteine, L-cysteine, and O-acetyl-L-serine to form alpha-keto acid, ammonia, and hydrogen sulfide or methanethiol, whereas activity toward cystathionine was negligible. These two isoenzymes showed notable differences in substrate specificity and pH optimum. In addition, we showed that EhMGL is an ideal target for the development of new chemotherapeutic agents against amebiasis by demonstrating an amebicidal effect of the methionine analog trifluoromethionine on trophozoites in culture (IC50 18 mum) and that this effect of trifluoromethionine was completely abolished by the addition of the MGL-specific inhibitor DL-propargylglycine.

Amino Acids, Sulfur↗
PubMed1998

Microsporidia, amitochondrial protists, possess a 70-kDa heat shock protein gene of mitochondrial evolutionary origin.

An intronless gene encoding a protein of 592 amino acid residues with similarity to 70-kDa heat shock proteins (HSP70s) has been cloned and sequenced from the amitochondrial protist Encephalitozoon cuniculi (phylum Microsporidia). Southern blot analyses show the presence of a single gene copy located on chromosome XI. The encoded protein exhibits an N-terminal hydrophobic leader sequence and two motifs shared by proteobacterial and mitochondrially expressed HSP70 homologs. Phylogenetic analysis using maximum likelihood and evolutionary distances place the E. cuniculi sequence in the cluster of mitochondrially expressed HSP70s, with a higher evolutionary rate than those of homologous sequences. Similar results were obtained after cloning a fragment of the homologous gene in the closely related species E. hellem. The presence of a nuclear targeting signal-like sequence supports a role of the Encephalitozoon HSP70 as a molecular chaperone of nuclear proteins. No evidence for cytosolic or endoplasmic reticulum forms of HSP70 was obtained through PCR amplification. These data suggest that Encephalitozoon species have evolved from an ancestor bearing mitochondria, which is in disagreement with the postulated presymbiotic origin of Microsporidia. The specific role and intracellular localization of the mitochondrial HSP70-like protein remain to be elucidated.

Amino Acid Sequence↗
PubMed2000

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.

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.

Amino Acid Sequence↗
PubMed1994

Carbohydrate metabolism and physiology of the parasitic protist Trichomonas vaginalis studied in chemostats.

The parasitic protist Trichomonas vaginalis was cultured in chemostats with glucose or maltose as carbon and energy source. The maximum growth rate was about six divisions per day independent of the substrate, and the apparent Km for glucose was 0.375 mM. While growing on maltose, the growth rate depended linearly on the maltose concentration, indicating that in contrast to glucose metabolism a diffusion step is rate-limiting to maltose metabolism. Cultures were examined over a wide range of growth rates under four conditions: utilizing glucose or maltose as carbon and energy source, with the carbon source rate-limiting or present in excess. Cell density, cellular protein and carbohydrate content as well as residual substrate concentration in the culture fluid were measured at each steady state. The protein content was constant at 100 pg protein per cell except when T. vaginalis was cultured under glucose limitation; in the latter case, slow-growing cells had less protein than cells grown at high rates. When growing under glucose limitation T. vaginalis metabolism changed to become more energy efficient at growth rates exceeding about half the maximum rate. The maintenance energy at the low growth rates accounted for approximately half of the total carbon consumption, which is high in comparison to other micro-organisms. At low growth rates the yield on maltose exceeded that on glucose, when expressed in terms of carbon equivalents. The yields on maltose and glucose were equal, but much lower, when the carbon source was not rate-limiting.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1999

Influence of growth conditions on RNA levels in relation to activity of core metabolic enzymes in the parasitic protists Trypanosoma brucei and Trichomonas vaginalis.

Levels of mRNAs encoding metabolic enzymes and their cellular activities were measured on continuous culture samples of the parasitic protists Trypanosoma brucei and Trichomonas vaginalis. The organisms were grown in chemostats at varying growth rates under glucose limitation or in the presence of excess glucose (EG), resulting in extensive adaptation of the cellular activities of glycolytic enzymes. rRNA and mRNA for beta-tubulin were monitored as controls. In Trypanosoma brucei levels of all RNAs showed a biphasic dependence on growth rate (= dilution rate D), with a sharp increase at higher D values. Cellular RNA levels of Trichomonas vaginalis rate-limited by glucose decreased slightly with increasing D. In EG-grown cells the opposite trend was observed. Equal levels for both carbon regimes were observed at intermediate D values. In both species the ratio between rRNA and mRNA encoding beta-tubulin was constant, independent of the carbon regime. mRNA encoding metabolic enzymes showed varying degrees of correlation with rRNA and beta-tubulin mRNA. In contrast, there was little to no correlation between mRNA levels and the activities of the enzymes they encode, even though only one of these is allosterically regulated. The data indicate that RNA levels in Trypanosoma brucei and Trichomonas vaginalis are determined by growth rate and in the latter species by the availability of the carbon and energy source. Rates of synthesis of metabolic enzymes are most likely regulated at the post-transcriptional level.

Animals↗
PubMed2006

Complete nucleotide sequence and genome organization of a single-stranded RNA virus infecting the marine fungoid protist Schizochytrium sp.

The complete nucleotide sequence of the genomic RNA of a marine fungoid protist-infecting virus (Schizochytrium single-stranded RNA virus; SssRNAV) has been determined. The viral RNA is single-stranded with a positive sense and is 9,018 nt in length [excluding the 3' poly(A) tail]. It contains two long open reading frames (ORFs), which are separated by an intergenic region of 92 nt. The 5' ORF (ORF1) is preceded by an untranslated leader sequence of 554 nt. The 3' large ORF (ORF2) and an additional ORF (ORF3) overlap ORF2 by 431 nt and are followed by an untranslated region of 70 nt [excluding the 3' poly(A) tail]. The deduced amino acid sequences of ORF1 and ORF2 products show similarity to non-structural and structural proteins of dicistroviruses, respectively. However, Northern blot analysis suggests that SssRNAV synthesizes subgenomic RNAs to translate ORF2 and ORF3, showing that the translation mechanism of downstream ORFs is distinct from that of dicistroviruses. Furthermore, although considerable similarities were detected by using a blast genome database search, phylogenetic analysis based on both the nucleotide and amino acid sequences of the putative RNA-dependent RNA polymerase (RdRp) and the RNA helicase suggests that SssRNAV is phylogenetically distinct from other virus families. Therefore, it is concluded that SssRNAV is not a member of any currently defined virus family and belongs to a novel, unrecognized virus group.

Amino Acid Sequence↗
PubMed2006

Prey selectivity of bacterivorous protists in different size fractions of reservoir water amended with nutrients.

An experiment designed to examine food preferences of heterotrophic nanoflagellates (HNF) grazing on bacterioplankton was performed in the freshwater Rímov reservoir (Czech Republic). Water samples were size-fractionated to obtain < 5 microm filtrate containing bacteria and HNF. To manipulate resource availability, < 5 microm treatments were incubated in dialysis bags submerged in the barrels filled with the unfiltered reservoir water amended with either orthophosphate or glucose or combination of both. We employed rRNA-targeted probes to assess HNF prey preferences by analysing bacterial prey in HNF food vacuoles compared with available bacteria. Actinobacteria (the HGC69a probe) were avoided by HNF in all treatments. Cytophaga-Flavobacterium-Bacteroidetes bacteria (the CF319a probe) were positively selected mainly in treatments in which bacteria were heavily grazed, the < 5 microm treatments, but this trend was less pronounced towards the end of the study. The members of a small subcluster of Betaproteobacteria (the R-BT065 probe) were mostly positively selected. The nutrient amendments differentially affected bacterioplankton dynamics in almost all treatments, and together with the size fractionation, altered HNF overall bacterivory as well as prey selection. Analyses of bacterivores in unfiltered treatments allowed to detect the effect of different protists on shifts in HNF selectivity observed in < 5 microm compared with unfiltered treatments.

Actinobacteria↗
PubMedDate not supplied

Purification and specificity of two alpha-glucosidase isoforms of the parasitic protist Trichomonas vaginalis.

Two isoforms of alpha-glucosidase were purified from the parasitic protist Trichomonas vaginalis. Both consisted of 103 kDa subunits, but differed in pH optimum and substrate specificity. Isoform 1 had a pH optimum around 4.5 and negligible activity on glucose oligomers other than maltose, while isoform 2 with a pH optimum of 5.5 hydrolyzed also such substrates at considerable rates. Neither had activity on glycogen or starch. Isoform 1 had a specific activity for hydrolysis of maltose of 30 U/mg protein and isoform 2 101 U/mg protein. The Km values were 0.4 mM and 2.0 mM, respectively. Isoform 2 probably corresponds to the activity detected on the cell surface.

Animals↗
PubMedDate not supplied

Contributions of the axostyle and flagella to closed mitosis in the protists Tritrichomonas foetus and Trichomonas vaginalis.

Tritrichomonas foetus and Trichomonas vaginalis are protists that undergo closed mitosis: the nuclear envelope remains intact and the spindle remains extranuclear. Here we show, in disagreement with previous studies, that the axostyle does not disappear during mitosis but rather actively participates in it. We document the main structural modifications of the cell during its cell cycle using video enhanced microscopy and computer animation, bright field light microscopy, confocal laser scanning microscopy, and scanning and transmission electron microscopy. We propose six phases in the trichomonad's cell cycle: an orthodox interphase, a pre-mitotic phase, and four stages during the cell division process. We report that in T. foetus and T. vaginalis: a) all skeletal structures such as the costa, pelta-axostyle system, basal bodies, flagella, and associated filaments of the mastigont system are duplicated in a pre-mitotic phase; b) the axostyle does not disappear during mitosis, otherwise playing a fundamental role in this process; c) axostyles participate in the changes in the cell shape, contortion of the anterior region of the cell, and karyokinesis; d) flagella are not under assembly during mitosis, as previously stated by others, but completely formed before it; and e) cytokinesis is powered in part by cell locomotion.

Animals↗
PubMedDate not supplied

The nucleariid amoebae: more protists at the animal-fungal boundary.

Nucleariid amoebae are naked amoebae, generally characterized by a spherical or sometimes flattened body with radiating filopodia. Most species preferentially consume algal prey or cyanobacteria. Phylogenetic analyses of the small-subunit rRNA coding regions from four nucleariid amoebae place these species near the origin of the animal-fungal divergence, together with the choanoflagellate-Corallochytrium and the ichthyosporean clades. The species Nuclearia delicatula, N. moebiusi, and N. simplex form a monophyletic group, while ATCC 30864, tentatively but possibly incorrectly assigned to Nuclearia sp., represents a separate line of descent. These nucleariids are unrelated to the lineage containing the testate filose amoebae (Testaceafilosia). Our findings expand the morphological and phylogenetic diversity of protists at the animal-fungal divergence.

Amoeba↗
PubMedDate not supplied

"Novel cytoskeletal proteins in protists": introductory remarks.

Protists provide the opportunity to integrate analyses from a low (molecular) to a high (organism) level of complexity within a broad evolutionary framework. The perpectives they offer in the cytoskeletal field are discussed with respect to emerging concepts of cellular biology.

Animals↗
PubMedDate not supplied

Why the world needs protists!

In this brief review, literature references are given to researches--involving diverse species of protists--that support the author's firm conviction that the biological world of today absolutely requires the presence of numerous of these generally small and unicelled organisms if it is to survive. Examples supplied come from areas within the field of protistology sensu lato as widely separated as basic phycological research on photosynthesis and protozoological/medical/biomedical investigations on malaria and other pathogens of human beings. Emphasis is primarily on the most relevant works of the past 10-15 years, although historically highly significant papers of older vintage require at least indirect--and occasionally direct--citation.

Animals↗
PubMedDate not supplied

Predicted secondary structure of the foraminiferal SSU 3' major domain reveals a molecular synapomorphy for granuloreticulosean protists.

The small subunit ribosomal RNA genes of foraminiferal protists are the largest and most divergent of any eukaryote. We demonstrate that this foraminiferal sequence alteration represents a substantial modification to the small subunit ribosomal RNA structure, including a large (up to 350 nt) novel helix in a very well-conserved portion of the head domain. This modification dates from the beginning of the foraminiferal radiation and, within modern orders, is partially conserved at the sequence level, suggesting that it is a functional part of the ribosome. The pattern of conservation makes it particularly useful for determining lower-taxon relationships in morphologically ambiguous allogromiid foraminifera.

Animals↗
PubMedDate not supplied

A java applet for exploring the new higher level classification of eukaryotes with emphasis on the taxonomy of protists.

We have converted the hierarchically organized new higher level classification of eukaryotes with emphasis on the taxonomy of protists proposed by Adl et al. into an interactive and dynamic Java applet. The current version of the applet can be accessed via http://phylogenetics.bioapps.biozentrum.uni-wuerzburg.de/etv. We use the layout from a Degree-of-Interest tree (DOITree) that effectively displays all the taxonomic information as well as the phylogenetic relationships described in the original article by Adl et al. The tree was made using the Prefuse Toolkit for interactive information visualization. All browsers capable of using Java applets will be able to view the tree. The applet is freely available for scientists, teachers, and students.

Animals↗
PubMed2000

Spatial distribution of protists in the presence of macroaggregates in a marine system.

The spatial distribution of marine protistan communities in the presence of organic macroaggregates, formed from natural seawater, was studied in several microcosm experiments. The presence of macroaggregates had two main effects. First, the size of the communities of bacteria, flagellates and ciliates increased, as these communities were three orders of magnitude higher in the aggregates than in the microcosm water. In addition, it brought the diversification on the niches accessible to planktonic microorganisms, as three phases were formed: water, aggregates and aggregate-water interphase. Some of the detected protistan taxa were only found in the water, and therefore they can be considered as truly free-swimming protists. Others quickly colonised the aggregates, and finally, some of them showed a preference for the aggregate-water interphase. We discuss this spatial structuring of the protistan community on the basis of their feeding strategies and structural and behavioural characteristics.

Journal Article↗
PubMed2000

Molecular phylogeny of methanogens associated with flagellated protists in the gut and with the gut epithelium of termites.

The molecular phylogeny of methanogenic archaea associated with the flagellated protist species Dinenympha and Microjoenia in the gut of termites, Reticulitermes speratus and Hodotermopsis sjoestedti, and those attached to the gut epithelium was examined based on PCR-amplified small-subunit ribosomal RNA genes. The sequences identified were classified into six groups within the genus Methanobrevibacter, including groups of yet uncharacterized novel species. Closely related methanogens were shared between Microjoenia and some Dinenympha cells in each termite. The methanogens harbored by the flagellates were phylogenetically different from the methanogens associated with the gut epithelium, suggesting that distinct methanogen species showed distinct spatial distributions in the termite gut.

Journal Article↗
PubMed1995

Amino acid consumption by the parasitic, amoeboid protists Entamoeba histolytica and E. invadens.

Amino acid consumption by Entamoeba histolytica and E. invadens has been measured in order to assess the possible roles of amino acids as energy substrates. Mixtures of amino acids enhanced the growth of the parasites in complex medium and their survival in simple medium. The consumption of several amino acids by the parasites suspended in simple media was greater when glucose was absent, suggesting that they may act as alternative energy sources. Under these conditions, asparagine was consumed extremely rapidly by E. histolytica in particular, and arginine, leucine and threonine were used greatly by both species. There was also a marked consumption of aspartate, but this occurred even when glucose was present. These five amino acids and phenylalanine were the ones consumed in greatest amounts during growth of E. histolytica in complex medium. Under the same growth conditions, E. invadens also used asparagine, arginine, leucine and threonine and in addition there was a large consumption of serine and especially glutamate. In contrast, the aspartate concentration in the complex medium increased and there was also a net increase in the concentration of some other amino acids. Alanine was produced by both species when the parasites were incubated in simple medium with glucose, and in greater amounts during growth in complex media, suggesting that it is an end product of energy metabolism. The findings provide support for the suggestion that energy generation through amino acid catabolism may be a characteristic feature of anaerobic parasitic protists.

Amino Acids↗