Limits...
Crystal structure of malaria parasite nucleosome assembly protein: distinct modes of protein localization and histone recognition.

Gill J, Yogavel M, Kumar A, Belrhali H, Jain SK, Rug M, Brown M, Maier AG, Sharma A - J. Biol. Chem. (2009)

Bottom Line: Expression of green fluorescent protein-tagged PfNapL confirmed its exclusive localization to the parasite cytoplasm.A detailed analysis of PfNapL structure suggests unique histone binding properties.The crucial structural differences observed between parasite and yeast NAPs shed light on possible new modes of histone recognition by nucleosome assembly proteins.

View Article: PubMed Central - PubMed

Affiliation: Structural and Computational Biology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Road, New Delhi 110067, India.

ABSTRACT
Nucleosome assembly proteins (NAPs) are histone chaperones that are essential for the transfer and incorporation of histones into nucleosomes. NAPs participate in assembly and disassembly of nucleosomes and in chromatin structure organization. Human malaria parasite Plasmodium falciparum contains two nucleosome assembly proteins termed PfNapL and PfNapS. To gain structural insights into the mechanism of NAPs, we have determined and analyzed the crystal structure of PfNapL at 2.3 A resolution. PfNapL, an ortholog of eukaryotic NAPs, is dimeric in nature and adopts a characteristic fold seen previously for yeast NAP-1 and Vps75 and for human SET/TAF-1b (beta)/INHAT. The PfNapL monomer is comprised of domain I, containing a dimerization alpha-helix, and a domain II, composed of alpha-helices and a beta-subdomain. Structural comparisons reveal that the "accessory domain," which is inserted between the domain I and domain II in yeast NAP-1 and other eukaryotic NAPs, is surprisingly absent in PfNapL. Expression of green fluorescent protein-tagged PfNapL confirmed its exclusive localization to the parasite cytoplasm. Attempts to disrupt the PfNapL gene were not successful, indicating its essential role for the malaria parasite. A detailed analysis of PfNapL structure suggests unique histone binding properties. The crucial structural differences observed between parasite and yeast NAPs shed light on possible new modes of histone recognition by nucleosome assembly proteins.

Show MeSH

Related in: MedlinePlus

The overall structure of PfNapL. a, domain diagram for the NAP/SET family of proteins. Histone chaperone proteins comprise a central domain and low complexity N and C termini. The inserted accessory domain in yNAP-1 (yellow) is highlighted, which is missing both in PfNapL and hSET (set to scale). The brown bar is not set to scale, since the N and C termini are variable. b, structure of the PfNapL dimer generated using 2-fold crystallographic symmetry operation. Each monomer of PfNapL contains a domain I composed of dimerization helix α2 and domain II composed of a β subdomain (four antiparallel β strands) and α-helices on the other side. c, structure-based sequence alignment of PfNapL, yNAP-1, and hSET. The structures share high structural homology, except for an additional accessory domain between domain I and domain II in yNAP-1 (foreground colored yellow). Residues contributing to dimer formation are colored green. Conserved hydrophobic motifs, predicted NES, and predicted NLS in PfNapL are underlined in purple and blue, respectively. Conserved residues from the hSET mutagenesis data are colored red and blue. d, sequence alignment of PfNapL and other eukaryotic members of the NAP family: yNAP-1 (Saccharomyces cerevisiae), xNAP-1 (Xenopus laevis), dNAP-1 (Drosophila melanogaster), and hNAP-1 (Homo sapiens). The accessory domain in yNAP-1 is foreground-colored yellow. The identical/conserved surface residues and conserved hydrophobic motifs are colored pink and purple, respectively. e, phylogenetic tree of NAPs from various species showing greater evolutionary distance of malaria parasite NAPs from homologs in yeast and humans (indicated by red arrows). f, model for in vitro relay of histones by P. falciparum nucleosome assembly proteins. This model was reproduced from Ref. 14, showing distinct roles for PfNapL (in cytoplasm as a histone carrier) and PfNapS (in nucleus capable of histone deposition). PfNapL (L) interacts with histones and can deliver histones to PfNapS (S; phosphorylated or unphosphorylated) readily. PfNapL upon phosphorylation (Lp) binds 3-fold better to the histones when compared with the unphosphorylated form of PfNapL. Phospho-PfNapL may deliver its histone cargo PfNapS. The latter takes over histones from Phospho-PfNapL-histone complexes and shuttles the histones into nucleus.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2665062&req=5

fig1: The overall structure of PfNapL. a, domain diagram for the NAP/SET family of proteins. Histone chaperone proteins comprise a central domain and low complexity N and C termini. The inserted accessory domain in yNAP-1 (yellow) is highlighted, which is missing both in PfNapL and hSET (set to scale). The brown bar is not set to scale, since the N and C termini are variable. b, structure of the PfNapL dimer generated using 2-fold crystallographic symmetry operation. Each monomer of PfNapL contains a domain I composed of dimerization helix α2 and domain II composed of a β subdomain (four antiparallel β strands) and α-helices on the other side. c, structure-based sequence alignment of PfNapL, yNAP-1, and hSET. The structures share high structural homology, except for an additional accessory domain between domain I and domain II in yNAP-1 (foreground colored yellow). Residues contributing to dimer formation are colored green. Conserved hydrophobic motifs, predicted NES, and predicted NLS in PfNapL are underlined in purple and blue, respectively. Conserved residues from the hSET mutagenesis data are colored red and blue. d, sequence alignment of PfNapL and other eukaryotic members of the NAP family: yNAP-1 (Saccharomyces cerevisiae), xNAP-1 (Xenopus laevis), dNAP-1 (Drosophila melanogaster), and hNAP-1 (Homo sapiens). The accessory domain in yNAP-1 is foreground-colored yellow. The identical/conserved surface residues and conserved hydrophobic motifs are colored pink and purple, respectively. e, phylogenetic tree of NAPs from various species showing greater evolutionary distance of malaria parasite NAPs from homologs in yeast and humans (indicated by red arrows). f, model for in vitro relay of histones by P. falciparum nucleosome assembly proteins. This model was reproduced from Ref. 14, showing distinct roles for PfNapL (in cytoplasm as a histone carrier) and PfNapS (in nucleus capable of histone deposition). PfNapL (L) interacts with histones and can deliver histones to PfNapS (S; phosphorylated or unphosphorylated) readily. PfNapL upon phosphorylation (Lp) binds 3-fold better to the histones when compared with the unphosphorylated form of PfNapL. Phospho-PfNapL may deliver its histone cargo PfNapS. The latter takes over histones from Phospho-PfNapL-histone complexes and shuttles the histones into nucleus.

Mentions: Malaria is one of the most common infectious diseases and remains an enormous public health problem. Malaria is caused by protozoan parasites of the genus Plasmodium, and the most serious forms of the disease are caused by Plasmodium falciparum (12). It is therefore crucial to identify, dissect, and exploit the molecular motors of malaria parasites, which can serve as essential targets for antimalarials. The human malaria parasite P. falciparum contains two nucleosome assembly proteins, which we have termed PfNapL and PfNapS and which are orthologs of eukaryotic NAPs (13). We have shown that both PfNapL and PfNapS are present in all erythrocytic stages of the parasite (13). PfNapL forms complexes with both histone tetramer and octamer and is predominantly localized in the cytoplasm in the asexual and sexual stages of the parasite (13, 14). PfNapL by itself is unable to deposit the histones onto DNA, but it can interact with both core and linker histones and is involved in histone binding, shuttling, and transfer/release, as shown earlier (13, 14) (Fig. 1f reproduced here from Ref. 14). The histone binding characteristics of PfNapL have been detailed previously, including its ability to transfer cytoplasmic histones on to PfNapS (which is localized both in cytoplasm and the nucleus and transfers histones into the nucleus for deposition) (14). A model for relay of histones from parasite cytoplasm to the nucleus has also been proposed by us previously (see Ref. 14 and specifically Fig. 1f). PfNapL preferentially interacts with the H3-H4 tetramer histones over H2A and H2B histones. PfNapL and PfNapS do not interact with each other (14). To address the structural basis of the nucleosome assembly activity in P. falciparum, we have determined and analyzed the crystal structure of PfNapL. Here, we detail the PfNapL structure and compare it with histone chaperones like yeast NAP-1 (yNAP-1), human SET/TAF-1b (β)/INHAT (hSET), Vps75, and Asf1-histone complex (15-21) in order to provide new insights into the mechanism of histone recognition.


Crystal structure of malaria parasite nucleosome assembly protein: distinct modes of protein localization and histone recognition.

Gill J, Yogavel M, Kumar A, Belrhali H, Jain SK, Rug M, Brown M, Maier AG, Sharma A - J. Biol. Chem. (2009)

The overall structure of PfNapL. a, domain diagram for the NAP/SET family of proteins. Histone chaperone proteins comprise a central domain and low complexity N and C termini. The inserted accessory domain in yNAP-1 (yellow) is highlighted, which is missing both in PfNapL and hSET (set to scale). The brown bar is not set to scale, since the N and C termini are variable. b, structure of the PfNapL dimer generated using 2-fold crystallographic symmetry operation. Each monomer of PfNapL contains a domain I composed of dimerization helix α2 and domain II composed of a β subdomain (four antiparallel β strands) and α-helices on the other side. c, structure-based sequence alignment of PfNapL, yNAP-1, and hSET. The structures share high structural homology, except for an additional accessory domain between domain I and domain II in yNAP-1 (foreground colored yellow). Residues contributing to dimer formation are colored green. Conserved hydrophobic motifs, predicted NES, and predicted NLS in PfNapL are underlined in purple and blue, respectively. Conserved residues from the hSET mutagenesis data are colored red and blue. d, sequence alignment of PfNapL and other eukaryotic members of the NAP family: yNAP-1 (Saccharomyces cerevisiae), xNAP-1 (Xenopus laevis), dNAP-1 (Drosophila melanogaster), and hNAP-1 (Homo sapiens). The accessory domain in yNAP-1 is foreground-colored yellow. The identical/conserved surface residues and conserved hydrophobic motifs are colored pink and purple, respectively. e, phylogenetic tree of NAPs from various species showing greater evolutionary distance of malaria parasite NAPs from homologs in yeast and humans (indicated by red arrows). f, model for in vitro relay of histones by P. falciparum nucleosome assembly proteins. This model was reproduced from Ref. 14, showing distinct roles for PfNapL (in cytoplasm as a histone carrier) and PfNapS (in nucleus capable of histone deposition). PfNapL (L) interacts with histones and can deliver histones to PfNapS (S; phosphorylated or unphosphorylated) readily. PfNapL upon phosphorylation (Lp) binds 3-fold better to the histones when compared with the unphosphorylated form of PfNapL. Phospho-PfNapL may deliver its histone cargo PfNapS. The latter takes over histones from Phospho-PfNapL-histone complexes and shuttles the histones into nucleus.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2665062&req=5

fig1: The overall structure of PfNapL. a, domain diagram for the NAP/SET family of proteins. Histone chaperone proteins comprise a central domain and low complexity N and C termini. The inserted accessory domain in yNAP-1 (yellow) is highlighted, which is missing both in PfNapL and hSET (set to scale). The brown bar is not set to scale, since the N and C termini are variable. b, structure of the PfNapL dimer generated using 2-fold crystallographic symmetry operation. Each monomer of PfNapL contains a domain I composed of dimerization helix α2 and domain II composed of a β subdomain (four antiparallel β strands) and α-helices on the other side. c, structure-based sequence alignment of PfNapL, yNAP-1, and hSET. The structures share high structural homology, except for an additional accessory domain between domain I and domain II in yNAP-1 (foreground colored yellow). Residues contributing to dimer formation are colored green. Conserved hydrophobic motifs, predicted NES, and predicted NLS in PfNapL are underlined in purple and blue, respectively. Conserved residues from the hSET mutagenesis data are colored red and blue. d, sequence alignment of PfNapL and other eukaryotic members of the NAP family: yNAP-1 (Saccharomyces cerevisiae), xNAP-1 (Xenopus laevis), dNAP-1 (Drosophila melanogaster), and hNAP-1 (Homo sapiens). The accessory domain in yNAP-1 is foreground-colored yellow. The identical/conserved surface residues and conserved hydrophobic motifs are colored pink and purple, respectively. e, phylogenetic tree of NAPs from various species showing greater evolutionary distance of malaria parasite NAPs from homologs in yeast and humans (indicated by red arrows). f, model for in vitro relay of histones by P. falciparum nucleosome assembly proteins. This model was reproduced from Ref. 14, showing distinct roles for PfNapL (in cytoplasm as a histone carrier) and PfNapS (in nucleus capable of histone deposition). PfNapL (L) interacts with histones and can deliver histones to PfNapS (S; phosphorylated or unphosphorylated) readily. PfNapL upon phosphorylation (Lp) binds 3-fold better to the histones when compared with the unphosphorylated form of PfNapL. Phospho-PfNapL may deliver its histone cargo PfNapS. The latter takes over histones from Phospho-PfNapL-histone complexes and shuttles the histones into nucleus.
Mentions: Malaria is one of the most common infectious diseases and remains an enormous public health problem. Malaria is caused by protozoan parasites of the genus Plasmodium, and the most serious forms of the disease are caused by Plasmodium falciparum (12). It is therefore crucial to identify, dissect, and exploit the molecular motors of malaria parasites, which can serve as essential targets for antimalarials. The human malaria parasite P. falciparum contains two nucleosome assembly proteins, which we have termed PfNapL and PfNapS and which are orthologs of eukaryotic NAPs (13). We have shown that both PfNapL and PfNapS are present in all erythrocytic stages of the parasite (13). PfNapL forms complexes with both histone tetramer and octamer and is predominantly localized in the cytoplasm in the asexual and sexual stages of the parasite (13, 14). PfNapL by itself is unable to deposit the histones onto DNA, but it can interact with both core and linker histones and is involved in histone binding, shuttling, and transfer/release, as shown earlier (13, 14) (Fig. 1f reproduced here from Ref. 14). The histone binding characteristics of PfNapL have been detailed previously, including its ability to transfer cytoplasmic histones on to PfNapS (which is localized both in cytoplasm and the nucleus and transfers histones into the nucleus for deposition) (14). A model for relay of histones from parasite cytoplasm to the nucleus has also been proposed by us previously (see Ref. 14 and specifically Fig. 1f). PfNapL preferentially interacts with the H3-H4 tetramer histones over H2A and H2B histones. PfNapL and PfNapS do not interact with each other (14). To address the structural basis of the nucleosome assembly activity in P. falciparum, we have determined and analyzed the crystal structure of PfNapL. Here, we detail the PfNapL structure and compare it with histone chaperones like yeast NAP-1 (yNAP-1), human SET/TAF-1b (β)/INHAT (hSET), Vps75, and Asf1-histone complex (15-21) in order to provide new insights into the mechanism of histone recognition.

Bottom Line: Expression of green fluorescent protein-tagged PfNapL confirmed its exclusive localization to the parasite cytoplasm.A detailed analysis of PfNapL structure suggests unique histone binding properties.The crucial structural differences observed between parasite and yeast NAPs shed light on possible new modes of histone recognition by nucleosome assembly proteins.

View Article: PubMed Central - PubMed

Affiliation: Structural and Computational Biology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Road, New Delhi 110067, India.

ABSTRACT
Nucleosome assembly proteins (NAPs) are histone chaperones that are essential for the transfer and incorporation of histones into nucleosomes. NAPs participate in assembly and disassembly of nucleosomes and in chromatin structure organization. Human malaria parasite Plasmodium falciparum contains two nucleosome assembly proteins termed PfNapL and PfNapS. To gain structural insights into the mechanism of NAPs, we have determined and analyzed the crystal structure of PfNapL at 2.3 A resolution. PfNapL, an ortholog of eukaryotic NAPs, is dimeric in nature and adopts a characteristic fold seen previously for yeast NAP-1 and Vps75 and for human SET/TAF-1b (beta)/INHAT. The PfNapL monomer is comprised of domain I, containing a dimerization alpha-helix, and a domain II, composed of alpha-helices and a beta-subdomain. Structural comparisons reveal that the "accessory domain," which is inserted between the domain I and domain II in yeast NAP-1 and other eukaryotic NAPs, is surprisingly absent in PfNapL. Expression of green fluorescent protein-tagged PfNapL confirmed its exclusive localization to the parasite cytoplasm. Attempts to disrupt the PfNapL gene were not successful, indicating its essential role for the malaria parasite. A detailed analysis of PfNapL structure suggests unique histone binding properties. The crucial structural differences observed between parasite and yeast NAPs shed light on possible new modes of histone recognition by nucleosome assembly proteins.

Show MeSH
Related in: MedlinePlus