Proteome Profiling of Leishmania Infantum Promastigotes

Abstract

A proteome analysis of the promastigote stage of the trypanosomatid parasite Leishmania infantum (MON-1 zymodeme) is described here for the first time. Total protein extracts were prepared at early logarithmic and stationary phases of replicate axenic cultures and processed by 2D electrophoresis (pH 3-10). A total of 28 differentially regulated proteins were identified by matrix-assisted laser desorption/ionization-tandem time of flight mass spectrometry. This approach has revealed that the electron transfer flavoprotein (ETF) and the eukaryotic elongation factor 1α (eEF1α) subunit have the same differential expression pattern at the protein and mRNA levels, up-regulation in the stationary phase. A low-molecular-weight isoform and an alternatively processed form of the eEF1α subunit have been detected. A 51 kDa subunit of replication factor A is up-regulated in dividing logarithmic promastigotes. None of the proteins described here shows opposite differential regulation values with the corresponding mRNA levels. Taken together with previous approaches to the proteome and the transcriptome, this report contributes to the elucidation of the differential regulation patterns of the ETF, the eEF1α subunit, the 40S ribosomal protein S12, α-tubulin and the T-complex protein 1 subunit γ throughout the life cycle of the parasites from the genus Leishmania.

Genomic Analysis of Sequence-Dependent DNA Curvature in Leishmania

“…Following the completion of the genome sequencing of the Tritryps (the three related trypanosomatid parasites: T. brucei, T. cruzi , and L. major ) [47], [48], [49], genome wide approaches in L. major have been mainly aimed at the characterization of global gene and protein expression profiles during development and in response to drugs, as well as protein subcellular localization, and host-parasite interactions [18], [50], [51], [52], [53], [54], [55], [56], [57], [58], [59]. While nucleic acid signals acting on different steps of the gene expression process have been described, search efforts have focused on primary sequence signals and no elements involved in transcription initiation of protein coding genes have been found [2].…”

Differential protein abundance in promastigotes of nitric oxide‐sensitive and resistant Leishmania chagasi strains

“…The spots of interest according to the previous analysis were excised with EXQuest Spot Cutter (BioRad), digested with trypsin and prepared for MALDI‐TOF/TOF MS as we described . A 0.8 μL aliquot of peptides from each spot was deposited in an OptiTOF™ Plate (Life Technologies) and a 0.8 μL drop of a 3 g/L α‐cyano‐4‐hydroxycinnamic solution (Sigma) was added to each one.…”

Proteomics and Leishmaniasis: Potential Clinical Applications

“…Another study using 2DE combined with MS evaluated the differential proteome expression of L. infantum promastigotes at exponential and stationary growth phases. A total of 28 differentially expressed proteins were identified and distinct proteoforms of eukaryotic elongation factor (EF) 1α were detected . Using a combination of 2DE coupled to MS/MS analysis of prefractionated proteins, a detailed view of stage‐specific proteoforms modulation in L. infantum promastigote and amastigote life stages was provided .…”

“…A total of 28 differentially expressed proteins were identified and distinct proteoforms of eukaryotic elongation factor (EF) 1α were detected. [52] Using a combination of 2DE coupled to MS/MS analysis of prefractionated proteins, a detailed view of stage-specific proteoforms modulation in L. infantum promastigote and amastigote life stages was provided. [53] Proteoforms of enolase, HSP70 related protein 1, Leishmania-activated C-kinase, eukaryotic initiation factor (eIF) 4a/5a and adenosyl homocysteinase were reported during L. donovani life stage differentiation from promastigotes to amastigotes [54] by 2DE coupled to MALDI-TOF and PMF.…”