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1.  Crystallization and preliminary X-ray crystallographic analysis of human autotaxin 
The α isoform of human autotaxin has been crystallized. Diffraction data were collected to 3.0 Å resolution using synchrotron radiation.
Autotaxin (ATX), which is also known as ectonucleotide pyrophosphatase/phosphodiesterase 2 (NPP2 or ENPP2) or phosphodiesterase Iα (PD-Iα), is an extracellular lysophospholipase D which generates lysophosphatidic acid (LPA) from lysophosphatidylcholine (LPC). ATX stimulates tumour-cell migration, angiogenesis and metastasis and is an attractive target for cancer therapy. For crystallographic studies, the α isoform of human ATX was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. X-ray diffraction data were collected to 3.0 Å resolution from a monoclinic crystal form belonging to space group C2, with unit-cell parameters a = 311.4, b = 147.9, c = 176.9 Å, β = 122.6°.
doi:10.1107/S174430911005311X
PMCID: PMC3080147  PMID: 21505238
autotaxin; lysophosphatidic acid; lysophospholipase D; ectonucleotide pyrophosphatase/phosphodiesterase 2
2.  Crystallization of mouse S-adenosyl-l-homocysteine hydrolase 
Mouse S-adenosyl-l-homocysteine hydrolase has been crystallized in the presence of the reaction product adenosine. Diffraction data to 1.55 Å resolution were collected using synchrotron radiation.
S-Adenosyl-l-homocysteine hydrolase (SAHH; EC 3.3.1.1) catalyzes the reversible hydrolysis of S-adenosyl-l-homocysteine to adenosine and l-homo­cysteine. For crystallographic investigations, mouse SAHH (MmSAHH) was overexpressed in bacterial cells and crystallized using the hanging-drop vapour-diffusion method in the presence of the reaction product adenosine. X-ray diffraction data to 1.55 Å resolution were collected from an orthorhombic crystal form belonging to space group I222 with unit-cell parameters a = 100.64, b = 104.44, c = 177.31 Å. Structural analysis by molecular replacement is in progress.
doi:10.1107/S1744309110000771
PMCID: PMC2833045  PMID: 20208169
S-adenosyl-l-homocysteine hydrolase; SAHH
3.  Crystallization and preliminary X-ray crystallographic study of 1-deoxy-d-xylulose 5-­phosphate reductoisomerase from Plasmodium falciparum  
1-Deoxy-d-xylulose 5-phosphate reductoisomerase from P. falciparum has been crystallized in the presence of NADPH. Diffraction data to 1.85 Å resolution have been collected using synchrotron radiation.
The nonmevalonate pathway of isoprenoid biosynthesis present in Plasmodium falciparum is known to be an effective target for antimalarial drugs. The second enzyme of the nonmevalonate pathway, 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR), catalyzes the transformation of 1-deoxy-d-xylulose 5-phosphate (DXP) to 2-C-methyl-d-erythritol 4-phosphate (MEP). For crystallographic studies, DXR from the human malaria parasite P. falciparum (PfDXR) was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method in the presence of NADPH. X-ray diffraction data to 1.85 Å resolution were collected from a monoclinic crystal form belonging to space group C2 with unit-cell parameters a = 168.89, b = 59.65, c = 86.58 Å, β = 117.8°. Structural analysis by molecular replacement is in progress.
doi:10.1107/S1744309110001739
PMCID: PMC2833050  PMID: 20208174
1-deoxy-d-xylulose 5-phosphate reductoisomerase; malaria; nonmevalonate pathway
4.  Crystallization and preliminary X-ray crystallographic study of phosphoglucose isomerase from Plasmodium falciparum  
Phosphoglucose isomerase from P. falciparum has been crystallized. Diffraction data to 1.8 Å resolution have been collected using synchrotron radiation.
Phosphoglucose isomerase (PGI) is a key enzyme in glycolysis and glycogenesis that catalyses the interconversion of glucose 6-phosphate (G6P) and fructose 6-­phosphate (F6P). For crystallographic studies, PGI from the human malaria parasite Plasmodium falciparum (PfPGI) was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. X-ray diffraction data to 1.5 Å resolution were collected from an orthorhombic crystal form belonging to space group P212121 with unit-cell parameters a = 103.3, b = 104.1, c = 114.6 Å. Structural analysis by molecular replacement is in progress.
doi:10.1107/S1744309110001740
PMCID: PMC2833051  PMID: 20208175
glucose 6-phosphate isomerase; malaria; phosphoglucose isomerase; phosphohexose isomerase
5.  Molecular basis of fosmidomycin's action on the human malaria parasite Plasmodium falciparum 
Scientific Reports  2011;1:9.
The human malaria parasite Plasmodium falciparum is responsible for the deaths of more than a million people each year. Fosmidomycin has been proven to be efficient in the treatment of P. falciparum malaria by inhibiting 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR), an enzyme of the non-mevalonate pathway, which is absent in humans. However, the structural details of DXR inhibition by fosmidomycin in P. falciparum are unknown. Here, we report the crystal structures of fosmidomycin-bound complete quaternary complexes of PfDXR. Our study revealed that (i) an intrinsic flexibility of the PfDXR molecule accounts for an induced-fit movement to accommodate the bound inhibitor in the active site and (ii) a cis arrangement of the oxygen atoms of the hydroxamate group of the bound inhibitor is essential for tight binding of the inhibitor to the active site metal. We expect the present structures to be useful guides for the design of more effective antimalarial compounds.
doi:10.1038/srep00009
PMCID: PMC3216497  PMID: 22355528
6.  Functional Complementation and Genetic Deletion Studies of KirBac Channels 
The Journal of Biological Chemistry  2010;285(52):40754-40761.
The superfamily of prokaryotic inwardly rectifying (KirBac) potassium channels is homologous to mammalian Kir channels. However, relatively little is known about their regulation or about their physiological role in vivo. In this study, we have used random mutagenesis and genetic complementation in K+-auxotrophic Escherichia coli and Saccharomyces cerevisiae to identify activatory mutations in a range of different KirBac channels. We also show that the KirBac6.1 gene (slr5078) is necessary for normal growth of the cyanobacterium Synechocystis PCC6803. Functional analysis and molecular dynamics simulations of selected activatory mutations identified regions within the slide helix, transmembrane helices, and C terminus that function as important regulators of KirBac channel activity, as well as a region close to the selectivity filter of KirBac3.1 that may have an effect on gating. In particular, the mutations identified in TM2 favor a model of KirBac channel gating in which opening of the pore at the helix-bundle crossing plays a far more important role than has recently been proposed.
doi:10.1074/jbc.M110.175687
PMCID: PMC3003375  PMID: 20876570
Ion Channels; Membrane Biophysics; Membrane Proteins; Potassium Channels; Potassium Transport; Kir Channel; KirBac
7.  Structural basis for recognition of cognate tRNA by tyrosyl-tRNA synthetase from three kingdoms 
Nucleic Acids Research  2007;35(13):4289-4300.
The specific aminoacylation of tRNA by tyrosyl-tRNA synthetases (TyrRSs) relies on the identity determinants in the cognate tRNATyrs. We have determined the crystal structure of Saccharomyces cerevisiae TyrRS (SceTyrRS) complexed with a Tyr-AMP analog and the native tRNATyr(GΨA). Structural information for TyrRS–tRNATyr complexes is now full-line for three kingdoms. Because the archaeal/eukaryotic TyrRSs–tRNATyrs pairs do not cross-react with their bacterial counterparts, the recognition modes of the identity determinants by the archaeal/eukaryotic TyrRSs were expected to be similar to each other but different from that by the bacterial TyrRSs. Interestingly, however, the tRNATyr recognition modes of SceTyrRS have both similarities and differences compared with those in the archaeal TyrRS: the recognition of the C1-G72 base pair by SceTyrRS is similar to that by the archaeal TyrRS, whereas the recognition of the A73 by SceTyrRS is different from that by the archaeal TyrRS but similar to that by the bacterial TyrRS. Thus, the lack of cross-reactivity between archaeal/eukaryotic and bacterial TyrRS-tRNATyr pairs most probably lies in the different sequence of the last base pair of the acceptor stem (C1-G72 vs G1-C72) of tRNATyr. On the other hand, the recognition mode of Tyr-AMP is conserved among the TyrRSs from the three kingdoms.
doi:10.1093/nar/gkm417
PMCID: PMC1934993  PMID: 17576676

Results 1-7 (7)