Identification of R2TP complex of Leishmania donovani and Plasmodium falciparum using genome wide in-silico analysis

Abstract

Recently discovered R2TP complex is an important multiprotein complex involved in multiple cellular process like snoRNP biogenesis, PIKK signaling, RNA polymerase II assembly and apoptosis. Within R2TP complex, Pih1 tightly interacts with Rvb1/Rvb2 and with Tah1 to form R2TP macromolecular complex. R2TP complex further interacts with Hsp90 to form R2TP-Hsp90 complex, which has been found critical in many cellular process. The genome wide screening of Leishmania donovani and Plasmodium falciparum led to the identification of RuvB like1, RuvB like 2, Pih1, and Tah1. Therefore, we speculate that this complex is also important for these parasites as in the yeast. The detailed analysis of crucial components of R2TP complex, Ld-RuvB like 1, and Ld-RuvB like 2, revealed the presence of characteristic motifs like DNA binding motif and ATPase motifs. Hsp90 is also reported from Leishmania donovani and Plasmodium falciparum suggesting that the R2TP complex further interacts with Hsp90 to form R2TP-Hsp90 complex. Recently it has been discovered that RuvB like proteins are overexpressed in many cancers and their ATPase activity is crucial for cancer cell proliferation and the human RuvBs have been proposed as suitable drug target for cancer. Similarly one of the Plasmodium falciparum RuvB like protein (PfRuvB3) has been found to be specific to the stage where nuclear division led multiplication of parasite take place. Considering all these it seems that the R2TP complex may be playing some critical role both in the cancer cell proliferation in human and rapid multiplication of the parasites Leishmania donovani and Plasmodium falciparum.

Keywords: AAA+ enzyme; ATPase; Pih1; Pontin; Reptin; RuvB like protein; Tah1.

Figure 1. Schematic diagram of R2TP complex involvement in diverse cellular activities. The schematic of R2TP complex involvement in various activities was prepared on the basis of reported role in yeast and human. R2TP complex is involved in the diverse cellular activities shown in the figure.

Figure 2. In-silico prediction of conserved motifs of RuvB from L. donovani using InterProScan. (A) Predicted motifs model of L. donovani RuvB1 was prepared, which shows that LdRuvB1 contains characteristic AAA+ domain and belongs to pontin (RuvB like1 helicase). (B) Conserved motifs model of L. donovani RuvB2 was prepared, which shows that Ld RuvB2 contains characteristic AAA+ domain and belongs to Reptin (RuvB like2 helicase).

Figure 3. Conserved motifs of L. donovani RuvB1 and comparison with yeast and human RuvB1 characteristic motifs. Multiple sequence alignment of LdRuvB1 with yeast and human RuvB1 using ClustalW at default parameters. The amino acid sequence of various conserved motifs like Walker A, Walker B, sensor I, arginine finger and sensor-II motifs are boxed in green color.

Figure 4. Conserved motifs of L. donovani RuvB2 and comparison with yeast and human RuvB2 characteristic motifs. Multiple sequence alignment of LdRuvB2 with yeast and human RuvB2 using ClustalW at default parameters. The amino acid sequence of various conserved motifs like Walker A, Walker B, sensor I, arginine finger, and sensor-II motifs are boxed in green color.

Figure 5. Structure modeling. The modeling of LdRuvB1 and LdRuvB2 was done using amino acid sequences at Swissmodel server. The molecular graphic images were produced using the UCSF Chimera package from the resource for Biocomputing, Visualization, and Informatics (http://www.cgl.ucsf.edu/chimera) at the University of California, San Francisco (supported by NIH P41 RR-01081). (A) (i) Template for LdRuvB1; (ii) LdRuvB1; (iii) superimposed image of template and LdRuvB1. (B) (i) Template for LdRuvB2; (ii) LdRuvB2; (iii) superimposed image of template and LdRuvB2. (C) Phylogenetic analysis of LdRuvB proteins with other RuvB proteins. The phylogenetic analysis of LdRuvB like proteins with E. coli, yeast and human RuvBs was carried by using Phylogeny.fr. with default parameters.

Figure 6. In-silico generated motifs model of Pih1 from L. donovani. (A) Predicted motifs model of LdPih1 was prepared with InterProScan which shows that LdPih1 contains characteristic PIH domain. (B) Predicted motifs model of PfPih1 was prepared with InterProScan, which shows characteristic PIH domain. (C) Phylogenetic analysis of LdPih1 and PfPih1 proteins with other Pih1 proteins was carried by using Phylogeny.fr.

Figure 7. Conserved motifs of Tah1 from L. donovani and P. falciparum. (A) Predicted motifs model of LdTah1 was prepared with InterProScan which shows that LdTah1 contain characteristic TPR domain. (B) Predicted motifs model of PfTah1 was prepared with InterProScan which shows characteristic TPR domain. (C) Phylogenetic analysis of LdTah1 and PfTah1 proteins with other Tah1 including yeast and human Tah1 proteins was carried by using Phylogeny.fr.

Figure 8. Structure modeling of Tah1 protein. The modeling of LdTah1 and PfTah1 was done using amino acid sequences at Swissmodel server. (A) Template, (B) LdTah1, (C) Superimposed image of template and LdTah1, (D) PfTah1, (E) Superimposed image of template and PfTah1. The molecular graphic images were produced using the UCSF Chimera package from the resource for Biocomputing, Visualization, and Informatics (http://www.cgl.ucsf.edu/chimera) at the University of California, San Francisco (supported by NIH P41 RR-01081).