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Journal of Molecular Cell Biology 12/2012;
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ABSTRACT: Aminoglycosides bind to the influenza A virus promoter (vRNA) at submicromolar concentration. The complex structure between the vRNA and neomycin illustrates that binding of neomycin causes a conformational change which would affect further transcription processes. Thus, aminoglycosides represent lead compounds for the discovery of antiviral therapeutics against influenza A virus.
Molecular BioSystems 09/2012; 8(11):2857-9. · 3.53 Impact Factor
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ABSTRACT: PA4608 is a single PilZ domain protein from Pseudomonas aeruginosa that binds to cyclic dimeric guanosine monophosphate (c-di-GMP). Although the monomeric structure of unbound PA4608 has been studied in detail, the molecular details of c-di-GMP binding to this protein are still uncharacterized. Hence, we determined the solution structure of c-di-GMP bound PA4608. We found that PA4608 undergoes conformational changes to expose the c-di-GMP binding site by ejection of the C-terminal 3(10) helix. A dislocation of the C-terminal tail in the presence of c-di-GMP implies that this region acts as a lid that alternately covers and exposes the hydrophobic surface of the binding site. In addition, mutagenesis and NOE data for PA4608 revealed that conserved residues are in contact with the c-di-GMP molecule. The unique structural characteristics of PA4608, including its monomeric state and its ligand binding characteristics, yield insight into its function as a c-di-GMP receptor.
Protein Science 02/2011; 20(2):270-7. · 2.80 Impact Factor
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ABSTRACT: Cyclic diguanylate (c-di-GMP) is a global regulator that modulates pathogen virulence and biofilm formation in bacteria. Although a bioinformatic study revealed that PilZ domain proteins are the long-sought c-di-GMP binding proteins, the mechanism by which c-di-GMP regulates them is uncertain. Pseudomonas putida PP4397 is one such protein that contains YcgR-N and PilZ domains and the apo-PP4397 structure was solved earlier by the Joint Center for Structural Genomics. We determined the crystal structure of holo-PP4397 and found that two intercalated c-di-GMPs fit into the junction of its YcgR-N and PilZ domains. Moreover, c-di-GMP binding induces PP4397 to undergo a dimer-to-monomer transition. Interestingly, another PilZ domain protein, VCA0042, binds to a single molecule of c-di-GMP, and both its apo and holo forms are dimeric. Mutational studies and the additional crystal structure of holo-VCA0042 (L135R) showed that the Arg122 residue of PP4397 is crucial for the recognition of two molecules of c-di-GMP. Thus, PilZ domain proteins exhibit different c-di-GMP binding stoichiometry and quaternary structure, and these differences are expected to play a role in generating diverse forms of c-di-GMP-mediated regulation.
Journal of Molecular Biology 03/2010; 398(1):97-110. · 4.00 Impact Factor
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ABSTRACT: Little attention has been paid to the specificity between E2 and the target protein during ubiquitination, although RING-E3 induces a potential intra-molecular reaction by mediating the direct transfer of ubiquitin from E2 to the target protein. We have constructed artificial E2 fusion proteins in which a target protein (p27) is tethered to one of six E2s via a flexible linker. Interestingly, only three E2s (UbcH5b, hHR6b, and Cdc34) are able to ubiquitinate p27 via an intra-molecular reaction in this system. Although the first ubiquitination of p27 (p27-Ub) by Cdc34 is less efficient than that of UbcH5b and hHR6b, the additional ubiquitin attachment to p27-Ub by Cdc34 is highly efficient. The E2 core of Cdc34 provides specificity to p27, and the residues 184-196 are required for possessive ubiquitination by Cdc34. We demonstrate direct E2 specificity for p27 and also show that differential ubiquitin linkages can be dependent on E2 alone.
BMB reports 01/2009; 41(12):852-7. · 1.72 Impact Factor
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ABSTRACT: Methylglyoxal (MG) is a toxic metabolite known to accumulate in various cell types. We detected in vivo conversion of MG to acetol in MG-accumulating Escherichia coli cells by use of (1)H nuclear magnetic resonance ((1)H-NMR) spectroscopy. A search for homologs of the mammalian aldo-keto reductases (AKRs), which are known to exhibit activity to MG, revealed nine open reading frames from the E. coli genome. Based on both sequence similarities and preliminary characterization with (1)H-NMR for crude extracts of the corresponding mutant strains, we chose five genes, yafB, yqhE, yeaE, yghZ, and yajO, for further study. Quantitative assessment of the metabolites produced in vitro from the crude extracts of these mutants and biochemical study with purified AKRs indicated that the yafB, yqhE, yeaE, and yghZ genes are involved in the conversion of MG to acetol in the presence of NADPH. When we assessed their in vivo catalytic activities by creating double mutants, all of these genes except for yqhE exhibited further sensitivities to MG in a glyoxalase-deficient strain. The results imply that the glutathione-independent detoxification of MG can occur through multiple pathways, consisting of yafB, yqhE, yeaE, and yghZ genes, leading to the generation of acetol.
Journal of Bacteriology 09/2005; 187(16):5782-9. · 3.83 Impact Factor
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Sun-Joo Lee,
So Jung Kim,
In-Kwon Kim, Junsang Ko,
Chang-Sook Jeong,
Gyung-Hwa Kim,
Chankyu Park,
Sa-Ouk Kang,
Pann-Ghill Suh,
Heung-Soo Lee,
Sun-Shin Cha
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ABSTRACT: Human DJ-1 and Escherichia coli Hsp31 belong to ThiJ/PfpI family, whose members contain a conserved domain. DJ-1 is associated with autosomal recessive early onset parkinsonism and Hsp31 is a molecular chaperone. Structural comparisons between DJ-1, Hsp31, and an Archaea protease, a member of ThiJ/PfpI family, lead to the identification of the chaperone activity of DJ-1 and the proteolytic activity of Hsp31. Moreover, the comparisons provide insights into how the functional diversity is realized in proteins that share an evolutionarily conserved domain. On the basis of the chaperone activity the possible role of DJ-1 in the pathogenesis of Parkinson's disease is discussed.
Journal of Biological Chemistry 12/2003; 278(45):44552-9. · 4.77 Impact Factor
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ABSTRACT: A yedU gene product with a molecular mass of 31 kDa is a hypothetical protein with no known function. The protein was purified and crystallized at 296 K. X-ray diffraction data have been collected to 2.3 A using synchrotron radiation. The crystals belong to the primitive orthorhombic system, with unit-cell parameters a = 50.56, b = 63.45, c = 168.02 A. The asymmetric unit contains two monomers of the protein, with a corresponding V(M) of 2.25 A(3) Da(-1) and a solvent content of 44.84%.
Acta Crystallographica Section D Biological Crystallography 08/2002; 58(Pt 7):1217-9. · 12.62 Impact Factor
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ABSTRACT: Conformational changes of periplasmic binding proteins are essential for their function in chemotaxis and transport. The allose-binding protein from Escherichia coli is, like other receptors in its family, composed of two alpha/beta domains joined by a three-stranded hinge. In the previously determined structure of the closed, ligand-bound form (Chaudhuri, B. N., Ko, J., Park, C., Jones, T. A., and Mowbray, S. L. (1999) J. Mol. Biol. 286, 1519-1531), the ligand-binding site is buried between the two domains. We report here the structures of three distinct open, ligand-free forms of this receptor, one solved at 3.1-A resolution and two others at 1.7-A resolution. Together, these allow a description of the conformational changes associated with ligand binding. A few large, coupled torsional changes in the hinge strands are sufficient to generate the overall bending motion, with only minor disruption of the individual domains. Integral water molecules appear to act as structural "ball bearings" in this process. The conformational changes of the related ribose-binding protein follow a distinct pattern. The observed differences between the two proteins can be interpreted in the context of changes in sequence and in crystal packing and provide new insights into the nature of hinge bending motion in this class of periplasmic binding proteins.
Journal of Biological Chemistry 05/2002; 277(16):14077-84. · 4.77 Impact Factor
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ABSTRACT: Conformational changes of periplasmic binding proteins are essential for their function in chemotaxis and transport. The allose-binding
protein from Escherichia coli is, like other receptors in its family, composed of two α/β domains joined by a three-stranded hinge. In the previously determined
structure of the closed, ligand-bound form (Chaudhuri, B. N., Ko, J., Park, C., Jones, T. A., and Mowbray, S. L. (1999) J. Mol. Biol. 286, 1519–1531), the ligand-binding site is buried between the two domains. We report here the structures of three distinct
open, ligand-free forms of this receptor, one solved at 3.1-Å resolution and two others at 1.7-Å resolution. Together, these
allow a description of the conformational changes associated with ligand binding. A few large, coupled torsional changes in
the hinge strands are sufficient to generate the overall bending motion, with only minor disruption of the individual domains.
Integral water molecules appear to act as structural “ball bearings” in this process. The conformational changes of the related
ribose-binding protein follow a distinct pattern. The observed differences between the two proteins can be interpreted in
the context of changes in sequence and in crystal packing and provide new insights into the nature of hinge bending motion
in this class of periplasmic binding proteins.
Journal of Biological Chemistry 04/2002; 277(16):14077-14084. · 4.77 Impact Factor
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ABSTRACT: ABC transport systems for import or export of nutrients and other substances across the cell membrane are widely distributed in nature. In most bacterial systems, a periplasmic component is the primary determinant of specificity of the transport complex as a whole. We report here the crystal structure of the periplasmic binding protein for the allose system (ALBP) from Escherichia coli, solved at 1.8 Å resolution using the molecular replacement method. As in the other members of the family (especially the ribose binding protein, RBP, with which it shares 35 % sequence homology), this structure consists of two similar domains joined by a three-stranded hinge region. The protein is believed to exist in a dynamic equilibrium of closed and open conformations in solution which is an important part of its function. In the closed ligand-bound form observed here, d-allose is buried at the domain interface. Only the β-anomer of allopyranose is seen in the crystal structure, although the α-anomer can potentially bind with a similar affinity. Details of the ligand-binding cleft reveal the features that determine substrate specificity. Extensive hydrogen bonding as well as hydrophobic interactions are found to be important. Altogether ten residues from both the domains form 14 hydrogen bonds with the sugar. In addition, three aromatic rings, one from each domain with faces parallel to the plane of the sugar ring and a third perpendicular, make up a hydrophobic stacking surface for the ring hydrogen atoms. Our results indicate that the aromatic rings forming the sugar binding cleft can sterically block the binding of any hexose epimer except d-allose, 6-deoxy-allose or 3-deoxy-glucose; the latter two are expected to bind with reduced affinity, due to the loss of some hydrogen bonds. The pyranose form of the pentose, d-ribose, can also fit into the ALBP binding cleft, although with lower binding affinity. Thus, ALBP can function as a low affinity transporter for d-ribose. The significance of these results is discussed in the context of the function of allose and ribose transport systems.
Journal of Molecular Biology 04/1999; · 4.00 Impact Factor
