The Structural Biology Center 19ID undulator beamline: facility specifications and protein crystallographic results

Rosenbaum, G.; Alkire, R. W.; Evans, Gwyndaf; Rotella, F. J.; Lazarski, K.; Zhang, Rong Guang; Ginell, Stephan L.; Duke, N.E.C.; Náday, I.; Lazarz, Jack; Molitsky, Michael; Keefe, Lisa J.; Gonczy, J. D.; Rock, Larry; Sanishvili, Ruslan; Walsh, Martin A.; Westbrook, Edwin M.; Joachimiak, Andrzej

doi:10.1107/s0909049505036721

Cited by 150 publications

(149 citation statements)

References 46 publications

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“…PA3455 was crystallized in the presence of 0.1 M Hepes-K (pH 7.0), 5% Tacsimate, and 10% PEG MME 5K, whereas the crystals of SMc02148 were grown in the presence of 0.1 M Bis-Tris (pH 7.5), 0.1 M Na-formate, 0.1 M Li-sulfate, and 0.3 M NDSB 211. Diffraction data were collected at the 19-ID beamline of the Structural Biology Center at the Advanced Photon Source (Argonne, IL) (29). The data were processed with HKL-3000 (30).…”

Section: Resultsmentioning

confidence: 99%

Polyphosphate-dependent synthesis of ATP and ADP by the family-2 polyphosphate kinases in bacteria

Nocek

Kochinyan

Proudfoot

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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168

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Inorganic polyphosphate (polyP) is a linear polymer of tens or hundreds of phosphate residues linked by high-energy bonds. It is found in all organisms and has been proposed to serve as an energy source in a pre-ATP world. This ubiquitous and abundant biopolymer plays numerous and vital roles in metabolism and regulation in prokaryotes and eukaryotes, but the underlying molecular mechanisms for most activities of polyP remain unknown. In prokaryotes, the synthesis and utilization of polyP are catalyzed by 2 families of polyP kinases, PPK1 and PPK2, and polyphosphatases. Here, we present structural and functional characterization of the PPK2 family. Proteins with a single PPK2 domain catalyze polyP-dependent phosphorylation of ADP to ATP, whereas proteins containing 2 fused PPK2 domains phosphorylate AMP to ADP. Crystal structures of 2 representative proteins, SMc02148 from Sinorhizobium meliloti and PA3455 from Pseudomonas aeruginosa, revealed a 3-layer ␣/␤/␣ sandwich fold with an ␣-helical lid similar to the structures of microbial thymidylate kinases, suggesting that these proteins share a common evolutionary origin and catalytic mechanism. Alanine replacement mutagenesis identified 9 conserved residues, which are required for activity and include the residues from both Walker A and B motifs and the lid. Thus, the PPK2s represent a molecular mechanism, which potentially allow bacteria to use polyP as an intracellular energy reserve for the generation of ATP and survival.crystal structure ͉ mutagenesis ͉ Walker motif ͉ AMP phosphorylation ͉ ADP phosphorylation I norganic polyphosphate (polyP) is a linear polymer composed of tens to hundreds of orthophosphate residues (P i ) linked by the energy-rich phosphoanhydride bonds, and it is found in all prokaryotes and eukaryotes (1-3). PolyP has numerous biological functions that include substitution for ATP in kinase reactions, acting as an energy source or storage reservoir of P i , chelating metals, and adjusting cellular physiology during growth, development, stress, starvation, and virulence (1, 4). Bacteria with low intracellular polyP levels show defective responses to various stress conditions, biofilm formation, quorum sensing, motility, and other virulence properties (3,5). Depending on the physiological state of the bacterium, the intracellular concentration of polyP is 0

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Section: Resultsmentioning

confidence: 99%

Polyphosphate-dependent synthesis of ATP and ADP by the family-2 polyphosphate kinases in bacteria

Nocek

Kochinyan

Proudfoot

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

113

168

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“…Diffraction data were collected at 100 K at the 19-ID beamline of the Structural Biology Center at the Advanced Photon Source, Argonne National Laboratory (38). The single wavelength anomalous dispersion (SAD) data at 0.97931 Å (12.6605 keV) up to 2.5 Å were collected using inverse-beam geometry near the selenium absorption edge from a single protein/DNA complex crystal (0.15 × 0.05 × 0.05 mm) of HetR and 21mer bearing a hetP promoter site.…”

Section: Methodsmentioning

confidence: 99%

Structures of complexes comprised of Fischerella transcription factor HetR with Anabaena DNA targets

Kim

Joachimiak

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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HetR is an essential regulator of heterocyst development in cyanobacteria. Many mutations in HetR render Anabaena incapable of nitrogen fixation. The protein binds to a DNA palindrome upstream of hetP and other genes. We have determined the crystal structures of HetR complexed with palindromic DNA targets, 21, 23, and 29 bp at 2.50-, 3.00-, and 3.25-Å resolution, respectively. The highest-resolution structure shows fine details of specific protein-DNA interactions. The lower-resolution structures with longer DNA duplexes have similar interaction patterns and show how the flap domains interact with DNA in a sequence nonspecific fashion. Fifteen of 15 protein-DNA contacts predicted on the basis of the structure were confirmed by single amino acid mutations that abolished binding in vitro and complementation in vivo. A striking feature of the structure is the association of glutamate 71 from each subunit of the HetR dimer with three successive cytosines in each arm of the palindromic target, a feature that is conserved among all known heterocyst-forming cyanobacteria sequenced to date.heterocyst differentiation | mutagenesis | X-ray crystallography

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“…In the case of the crystals from N. europea, the cryosolution was made of a 7:3 mixture of well solution and PEG400. Data collection for both proteins was done at beamline 19-ID of the Structural Biology Center (Rosenbaum et al 2006) at the Advanced Photon Source (APS). Data for both structures were collected at 100 K. Data collection, structure determination, and refinement statistics are summarized in Table 1.…”

Section: Data Collection Structure Determination and Refinementmentioning

confidence: 99%

Crystal structures of TM0549 and NE1324—two orthologs of E. coli AHAS isozyme III small regulatory subunit

et al. 2007

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Crystal structures of two orthologs of the regulatory subunit of acetohydroxyacid synthase III (AHAS, EC 2.2.1.6) from Thermotoga maritima (TM0549) and Nitrosomonas europea (NE1324) were determined by single-wavelength anomalous diffraction methods with the use of selenomethionine derivatives at 2.3 Å and 2.5 Å , respectively. TM0549 and NE1324 share the same fold, and in both proteins the polypeptide chain contains two separate domains of a similar size. Each protein contains a C-terminal domain with ferredoxin-type fold and an N-terminal ACT domain, of which the latter is characteristic for several proteins involved in amino acid metabolism. The ferredoxin domain is stabilized by a calcium ion in the crystal structure of NE1324 and by a Mg(H 2 O) 6 2+ ion in TM0549. Both TM0549 and NE1324 form dimeric assemblies in the crystal lattice.Keywords: acetohydroxyacid synthase; actolactate synthase; regulatory subunit; ACT domain; AHAS; protein refolding Acetohydroxyacid synthase (AHAS, EC 2.2.1.6) catalyzes two physiologically significant reactions in the synthesis of isoleucine, valine, and leucine. In the first reaction, which is an initial step in the synthesis of isoleucine, 2-aceto-2-hydroxy butyrate is produced by condensation of 2-ketobutyrate with pyruvate. In the second reaction, 2-acetolactate is synthesized from two pyruvate molecules, and this provides substrates for synthesis of valine and leucine (Umbarger 1978(Umbarger , 1987Chipman et al. 1998). This pathway of branched-chain amino acid biosynthesis is characteristic for bacteria, fungi, algae, and higher plants, but not for animals. Accordingly, AHAS inhibitors have been developed as herbicides which have found broad application (Short and Colborn 1999). The inhibitors of branched-chain amino acids synthesis are also being tested as potential antituberculosis agents (Grandoni et al. 1998;Zohar et al. 2003;Choi et al. 2005).Three different FAD-dependent AHAS isozymes (I, II, and III) are found in enterobacteria (e.g., Escherichia ), but most other organisms from the bacteria encode only a single AHAS enzyme, similar to isozyme III from E. coli. The acetohydroxyacid synthases are multimeric proteins, and most frequently their biological unit is composed of two catalytic subunits (CSU) and two small regulatory subunits (SSU). In the absence of the SSU subunit, the CSU dimer is unstable (Vyazmensky et al. 1996), suggesting that in the holoenzyme the AHAS dimer is stabilized by a SSU dimer. The large catalytic subunits also show very weak activity without their associated regulatory subunits (Weinstock et al. 1992;Pang and Duggleby 1999). The E. coli AHAS enzyme can be reconstituted with SSU subunits from other organisms (Porat et al. 2004), and the reconstituted enzymes form stable heterotetramers. The properly associated holoenzymes show full catalytic activity, and are also susceptible to valine inhibition.Previous mutagenesis studies (Mendel et al. 2001;Kaplun et al. 2006) revealed that the valine binding sites are located at the dimerizatio...

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The Structural Biology Center 19ID undulator beamline: facility specifications and protein crystallographic results

Cited by 150 publications

References 46 publications

Polyphosphate-dependent synthesis of ATP and ADP by the family-2 polyphosphate kinases in bacteria

Polyphosphate-dependent synthesis of ATP and ADP by the family-2 polyphosphate kinases in bacteria

Structures of complexes comprised of Fischerella transcription factor HetR with Anabaena DNA targets

Crystal structures of TM0549 and NE1324—two orthologs of E. coli AHAS isozyme III small regulatory subunit

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