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author:("Tanabe, miki")
1.  Crystal Structures of Acetate Kinases from the Eukaryotic Pathogens Entamoeba histolytica and Cryptococcus neoformans 
Journal of structural biology  2012;181(2):185-189.
Acetate kinases (ACKs) are members of the acetate and sugar kinase/hsp70/actin (ASKHA) superfamily and catalyze the reversible phosphorylation of acetate, with ADP/ATP the most common phosphoryl acceptor/donor. While prokaryotic ACKs have been the subject of extensive biochemical and structural characterization, there is a comparative paucity of information on eukaryotic ACKs, and prior to this report, no structure of an ACK of eukaryotic origin was available. We determined the structures of ACKs from the eukaryotic pathogens Entamoeba histolytica and Cryptococcus neoformans. Each active site is located at an interdomain interface, and the acetate and phosphate binding pockets display sequence and structural conservation with their prokaryotic counterparts. Interestingly, the E. histolytica ACK has previously been shown to be pyrophosphate (PPi)-dependent, and is the first ACK demonstrated to have this property. Examination of its structure demonstrates how subtle amino acid substitutions within the active site have converted cosubstrate specificity from ATP to PPi while retaining a similar backbone conformation. Differences in the angle between domains surrounding the active site suggest that interdomain movement may accompany catalysis. Taken together, these structures are consistent with the eukaryotic ACKs following a similar reaction mechanism as is proposed for the prokaryotic homologs.
doi:10.1016/j.jsb.2012.11.001
PMCID: PMC3565045  PMID: 23159802
Acetate kinase; PPi-dependent kinase; ASKHA superfamily
2.  Expression, purification and preliminary X-ray analysis of the Neisseria meningitidis outer membrane protein PorB 
The major outer membrane protein PorB from N. meningitidis was crystallized in three crystal forms; the best X-ray diffraction data were collected to 2.3 Å resolution.
The Neisseria meningitidis outer membrane protein PorB was expressed in Escherichia coli and purified from inclusion bodies by denaturation in urea followed by refolding in buffered LDAO on a size-exclusion column. PorB has been crystallized in three different crystal forms: C222, R32 and P63. The C222 crystal form may contain either one or two PorB monomers in the asymmetric unit, while both the R32 and P63 crystal forms contained one PorB monomer in the asymmetric unit. Of the three, the P63 crystal form had the best diffraction quality, yielding data extending to 2.3 Å resolution.
doi:10.1107/S1744309109032333
PMCID: PMC2765884  PMID: 19851005
outer membrane proteins; Neisseria meningitidis; denaturation; refolding; detergents; β-barrel membrane proteins; porins
3.  Expression, purification and preliminary X-ray analysis of the Neisseria meningitidis outer membrane protein PorB 
The Neisseria meningitidis outer membrane protein PorB was expressed in Escherichia coli and purified from inclusion bodies by denaturation in urea followed by refolding in buffered LDAO on a size exclusion column. PorB has been crystallized in three different crystal forms: C222, R32 and P63. The C222 crystal form may contain either one or two PorB monomers in the asymmetric unit while both the R32 and P63 crystal forms contain one PorB monomer in the asymmetric unit. Of the three, the P63 crystal form had the best diffraction quality, yielding data extending to 2.3 Å resolution.
doi:10.1107/S1744309109032333
PMCID: PMC2765884  PMID: 19851005
outer membrane protein; Neisseria meningitides; denaturation; refolding; detergent; β-barrel membrane protein; porin
4.  The ABC Transporter Protein OppA Provides Protection against Experimental Yersinia pestis Infection  
Infection and Immunity  2006;74(6):3687-3691.
The identification of Yersinia pestis as a potential bioterrorism agent and the emergence of antibiotic-resistant strains have highlighted the need for improved vaccines and treatments for plague. The aim of this study was to evaluate the potential for ATP-binding cassette (ABC) transporter proteins to be exploited as novel vaccines against plague. Western blotting of ABC transporter proteins using sera from rabbits immunized with killed whole Y. pestis cells or human convalescent-phase sera identified four immunologically reactive proteins: OppA, PstS, YrbD, and PiuA. Mice immunized with these proteins developed antibody to the immunogen. When the immunized mice were challenged with Y. pestis, the OppA-immunized mice showed an increased time to death compared to other groups, and protection appeared to correlate with the level of immunoglobulin G antibody to OppA.
doi:10.1128/IAI.01837-05
PMCID: PMC1479284  PMID: 16714605

Results 1-4 (4)