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Characterization and binding affinities of SmLANP: a new Schistosoma mansoni member of the ANP32 family of regulatory proteins.
Molecular and Biochemical Parasitology
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Members of the leucine-rich repeat protein family are involved in diverse functions including protein phosphatase 2-inhibition, cell cycle regulation, gene regulation and signalling pathways. A novel Schistosoma mansoni gene, called SmLANP, presenting homology to various genes coding for proteins that belong to the super family of leucine-rich repeat proteins, was characterized here. SmLANP was 1184bp in length as determined from cDNA and genomic sequences and encoded a 296 amino acid open reading frame that spanning from 6 to 894bp. The predicted amino acid sequence had a calculated molecular weight of 32kDa. Analysis of the predicted sequence indicated the presence of 3 leucine-rich domains (LRR) located in the N-terminal region and an aspartic acid rich region in the C-terminal end. SmLANP transcript is expressed in all stages of the S. mansoni life cycle analyzed, exhibiting the highest expression level in males. The SmLANP protein was expressed in a GST expression system and antibodies raised in mice against the recombinant protein. By immunolocalization assay, using adult worms, it was shown that the protein is mainly present in the cell nucleus through the whole body and strongly expressed along the tegument cell body nuclei of adult worms. As members of this family are usually involved in protein-protein interaction, a yeast two hybrid assay was conducted to identify putative binding partners for SmLANP. Thirty-six possible partners were identified, and a protein ATP synthase subunit alpha was confirmed by pull down assays, as a binding partner of the SmLANP protein.
Amino Acid Sequence, Animals, Antibodies, Helminth/metabolism, Gene Expression Regulation, Helminth Proteins/chemistry, Helminth Proteins/genetics, Immunohistochemistry, Intracellular Signaling Peptides and Proteins/chemistry, Intracellular Signaling Peptides and Proteins/genetics, Life Cycle Stages, Models, Molecular, Molecular Sequence Data, Protein Binding, Protein Structure, Tertiary, Recombinant Proteins/metabolism, Schistosoma mansoni/chemistry, Schistosoma mansoni/genetics, Sequence Alignment, Two-Hybrid System Techniques
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