Overlooked post-translational modifications of proteins in Plasmodium falciparum: N- and O-glycosylation - A Review

Human malignant malaria is caused by Plasmodium falciparum and accounts for almost 900,000 deaths per year, the majority of which are children and pregnant women in developing countries. There has been significant effort to understand the biology of P. falciparum and its interactions with the host. However, these studies are hindered because several aspects of parasite biology remain controversial, such as N- and O-glycosylation. This review describes work that has been done to elucidate protein glycosylation in P. falciparum and it focuses on describing biochemical evidence for N- and O-glycosylation. Although there has been significant work in this field, these aspects of parasite biochemistry need to be explored further.

á-N-acetylglucosamine-linked O-glycans of sialoglycoproteins (Tc-mucins) from Trypanosoma cruzi Colombiana strain

Trypanosoma cruzi sialoglycoproteins (Tc-mucins) are mucin-like molecules linked to a parasite membrane via a glycosylphosphatidylinositol anchor. We previously determined the structures of Tc-mucin O-glycan domains from several T. cruzi strains and observed significant differences among them. We now report the amino acid content and structure of Tc-mucin O-glycan chains from T. cruzi Colombiana, a strain resistant to common trypanocidal drugs. Amino acid analysis demonstrated the predominance of threonine residues (42%) and helped to identify the O-glycans as belonging to a Tc-mucin family that contain a ²-galactofuranose (²-Galf) residue attached to an á-N-acetylglucosamine (á-GlcNAc) O-4, with the most complex glycan, a pentasaccharide-GlcNAc-ol with a branched trigalactopyranose chain, on the GlcNAc O-6. The presence of ²-Galf on O-glycans from T. cruzi Colombiana mucins supports the use of glycosylation as a phylogenetic marker for the classification of Colombiana in the T. cruzi I group.

Endophytic colonization of rice (Oryza sativa L.) by the diazotrophic bacterium Burkholderia kururiensis and its ability to enhance plant growth

Burkholderia kururiensis is a diazotrophic bacterium originally isolated from a polluted aquifer environment and presents a high level of similarity with the rice endophyte "B. brasilensis" species. This work assessed the ability of B. kururiensis to endophytically colonize rice plantlets by monitoring different tissues of root-inoculated plants for the presence of bacterial growth in different media, electron microscopy and by 16S rDNA analysis. Observations of roots, stems and leaves of inoculated rice plantlets by electron microscopy revealed B. kururiensis colonization predominantly on root hair zones, demonstrating endophytic colonization primarily through the endodermis, followed by spreading into xylem vessels, a possible pathway leading to aerial parts. Although indifferent for the bacterial growth itself, addition of a nitrogen source was a limiting factor for endophytic colonization. As endophytic colonization was directly associated to an enhanced plant development, production of phytohormone auxin/indole-3-acetic acid by B. kururiensis was assayed with transgenic rice plantlets containing an auxin-responsive reporter (DR5-GUS). Our findings suggest the ability of auxin production by plant-associated B. kururiensis which may have a stimulatory effect on plant development, as evidenced by activation of DR5-GUS. We hereby demonstrate, for the first time, the ability of B. kururiensis to endophytically colonize rice, promoting both plant growth and rice grain yield.

Burkholderia kururiensis é uma bactéria diazotrófica, originalmente isolada de um ambiente aquático poluído e apresenta alto nível de similaridade com a espécie endofítica "B. brasilensis" encontrada na planta de arroz. Este artigo demonstrou a habilidade de B. kururiensis colonizar endofiticamente plântulas de arroz, após esta bactéria ter sido inoculada na raiz das plantas. Esta capacidade foi confirmada pelo crescimento bacteriano em diferentes tecidos da planta, por microscopia eletrônica e pela análise do 16S rADN. Observação por microscopia eletrônica das raízes, caule e folhas das plântulas de arroz inoculadas, revelou predominância da colonização de B. kururiensis na zona pilífera da raiz, demonstrando que a colonização endofítica inicia-se na endoderme, espalha-se pelo xilema, sendo esta a possível via para a bactéria alcançar as partes aéreas. A adição de uma fonte de nitrogênio, embora não tenha influenciado no crescimento bacteriano, foi um fator limitante para a colonização endofítica. Como a colonização endofítica mostrou-se diretamente associada ao aumento no desenvolvimento da planta, a produção do fitohormônio auxina/ácido 3-indolacético pela B. kururiensis foi verificada utilizando uma plântula de arroz transgênica, contendo o repórter responsivo para auxina (DR5-GUS). Nossos resultados sugerem que a produção de auxina pela B. kururiensis é responsável pelo estímulo no desenvolvimento da planta verificado pela ativação do DR5-GUS. Neste trabalho demonstramos, pela primeira vez, a habilidade de B. kururiensis colonizar endofiticamente a planta de arroz, promovendo tanto o aumento no crescimento da planta como a produção de sementes de arroz.

Strategies for the structure determination of parasite glycoconjugates using fast atom bombardment mass spectrometry

Fast atom bombardment mass spectrometry (FAB MS) is a sensitive and powerful technique for the analysis of thermally labile and involatile materials. In this paper we describe the application of the method to the characterization of oligosaccharides isolated from the cell surface glycoconjugates of parasitic protozoa. The spectra typically contain both molecular ions, which define the monosaccharide composition, and fragment ions which are related to the structure of the intact molecules. The use of additional techniques such as chemical derivatization and colisional activation enhances fragmentation and simplifies interpretation of the data, enabling the determination of residue sequence, the positions of branch points, and the location of noncarbohydrate substituents. We have applied these techniques to the characterization of phosphoinositol oligosaccharides from members of the Trypanosoma family, including Leptomonas smueli, Endotrypanum schaudinni and Leishmania adleri. Although it is not usually possible to determine the complete structure of oligosaccharide by mass spectrometric methods alone, the information gained greatly simplifies the interpretation of the results from other techiniques such as nuclear magnetic resonance and methylation analysis.

Evidence for the presence of ß-D-galactofuranosyl residues (1->3)-linked to a-D-mannopyranose on the cell membrane in all developmental stages of Trypanosoma cruzi

The lipopeptidophosphoglycan (LPPG) component isolated from epimastigote forms of Trypanosoma cruzi has antigenic properties. Using a rabbit antiserum aginst LPPG, specific for ß-D-galactofuranosyl residues (1-3)-linked to alfa-Dmannopyranose, we identified, by indirect immunofluorescence, the presence of epitopes containing ß-D-galactofuranosyl structures on the cell membrane of epimastigote, amastigote and trypomastigote forms of the parasite. The results demonstrade that all developmental stages of T. cruzi contain similar antigenic determinants on their cell body and flagellar membrane