Abstract
BackgroundProtein Phosphatase 1 (PP1) is an enzyme essential to cell viability in the malaria parasite Plasmodium falciparum (Pf). The activity of PP1 is regulated by the binding of regulatory subunits, of which there are up to 200 in humans, but only 3 have been so far reported for the parasite. To better understand the P. falciparum PP1 (PfPP1) regulatory network, we here report the use of three strategies to characterize the PfPP1 interactome: co-affinity purified proteins identified by mass spectrometry, yeast two-hybrid (Y2H) screening and in silico analysis of the P. falciparum predicted proteome.ResultsCo-affinity purification followed by MS analysis identified 6 PfPP1 interacting proteins (Pips) of which 3 contained the RVxF consensus binding, 2 with a Fxx[RK]x[RK] motif, also shown to be a PP1 binding motif and one with both binding motifs. The Y2H screens identified 134 proteins of which 30 present the RVxF binding motif and 20 have the Fxx[RK]x[RK] binding motif. The in silico screen of the Pf predicted proteome using a consensus RVxF motif as template revealed the presence of 55 potential Pips. As further demonstration, 35 candidate proteins were validated as PfPP1 interacting proteins in an ELISA-based assay.ConclusionsTo the best of our knowledge, this is the first study on PfPP1 interactome. The data reports several conserved PP1 interacting proteins as well as a high number of specific interactors to PfPP1. Their analysis indicates a high diversity of biological functions for PP1 in Plasmodium. Based on the present data and on an earlier study of the Pf interactome, a potential implication of Pips in protein folding/proteolysis, transcription and pathogenicity networks is proposed. The present work provides a starting point for further studies on the structural basis of these interactions and their functions in P. falciparum.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2571-z) contains supplementary material, which is available to authorized users.
Highlights
Protein Phosphatase 1 (PP1) is an enzyme essential to cell viability in the malaria parasite Plasmodium falciparum (Pf)
We showed that P. falciparum PP1 is submitted to a control of its activity by Pf Leucine Rich Repeat 1 (PfLRR1), Inhibitor-2 (PfI2) and -3 (PfI3) with substantial differences compared to the human orthologs for Inhibitor 2 and 3 [13,14,15,16,17,18]
Identification of P. falciparum PP1 (PfPP1) interacting proteins (Pips) by affinity/Mass Spectrometry In order to identify Pips expressed by P. falciparum, affinity purification on PfPP1 followed by mass spectrometry was carried out
Summary
Protein Phosphatase 1 (PP1) is an enzyme essential to cell viability in the malaria parasite Plasmodium falciparum (Pf). In Plasmodium falciparum (Pf), the most deadly apicomplexan parasite, efforts have begun for an examination of the biological roles of protein kinases and phosphatases and their potential as drug targets. In this context, biochemical and cloning studies have reported enzymatic activities and the genes of catalytic subunits related to plasmodial phosphatases. Biochemical and cloning studies have reported enzymatic activities and the genes of catalytic subunits related to plasmodial phosphatases These include PfPP1c, PfPP2A, Hollin et al BMC Genomics (2016) 17:246
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