scholarly journals Cryo-EM structure of a thermostable bacterial nanocompartment

IUCrJ ◽  
2021 ◽  
Vol 8 (3) ◽  
Author(s):  
Timothy Wiryaman ◽  
Navtej Toor
Keyword(s):  
High Resolution ◽  
Iron Metabolism ◽  
Thermotoga Maritima ◽  
Thermophilic Bacterium ◽  
Threefold Axis ◽  
Direct Role ◽  
Dimer Interface ◽  
Loop Domain

Protein nanocompartments are widespread in bacteria and archaea, but their functions are not yet well understood. Here, the cryo-EM structure of a nanocompartment from the thermophilic bacterium Thermotoga maritima is reported at 2.0 Å resolution. The high resolution of this structure shows that interactions in the E-loop domain may be important for the thermostability of the nanocompartment assembly. Also, the channels at the fivefold axis, threefold axis and dimer interface are assessed for their ability to transport iron. Finally, an unexpected flavin ligand was identified on the exterior of the shell, indicating that this nanocompartment may also play a direct role in iron metabolism.

scholarly journals High Resolution Crystal Structure of the Catalytic Domain of ADAMTS-5 (Aggrecanase-2)

2007 ◽  
Vol 283 (3) ◽  
pp. 1501-1507 ◽  
Author(s):  
Huey-Sheng Shieh ◽  
Karl J. Mathis ◽  
Jennifer M. Williams ◽  
Robert L. Hills ◽  
Joe F. Wiese ◽  
...  
Keyword(s):  
Crystal Structure ◽  
High Resolution ◽  
Catalytic Domain ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Direct Role ◽  
Three Dimensional Structure ◽  
High Resolution Structure ◽  
A Disintegrin And Metalloproteinase ◽  
Resolution Structure

Aggrecanase-2 (a disintegrin and metalloproteinase with thrombospondin motifs-5 (ADAMTS-5)), a member of the ADAMTS protein family, is critically involved in arthritic diseases because of its direct role in cleaving the cartilage component aggrecan. The catalytic domain of aggrecanase-2 has been refolded, purified, and crystallized, and its three-dimensional structure determined to 1.4Å resolution in the presence of an inhibitor. A high resolution structure of an ADAMTS/aggrecanase protein provides an opportunity for the development of therapeutics to treat osteoarthritis.

Cytoskeletal control of neuronal geometry: A serial EM analysis

1988 ◽  
Vol 46 ◽  
pp. 24-25
Author(s):  
John K. Stevens ◽  
Judy Trogadis
Keyword(s):  
High Resolution ◽  
High Speed ◽  
Three Dimensional ◽  
Video Camera ◽  
Electron Microscopic Level ◽  
Electron Microscopic ◽  
Direct Role ◽  
Live Image ◽  
Neuronal Geometry ◽  
Rotation Stage

The cytoskeleton plays a direct role in controlling neurite shape. To quantitatively study both the three dimensional shape and the sub-micron structure of the cytoskeleton requires complete serial reconstruction at the Electron Microscopic level. We have devised a computer reconstruction system specifically for this purpose.The system uses a 35mm film copy of 3.25 x 4.00 inch EM negative as the data source. The film is placed into a high speed film transport (15 frames/second), which is mounted on a X,Y and rotation stage controlled by stepping motors. The 35mm film is viewed through a stepping motor controlled zoom lens mounted on a high resolution (1119 x 1024) video camera. A high resolution frame grabber controlled by the computer can store one complete frame. Thus, the live image and a stored image may be displayed alternately on a high resolution monitor. Finally, a graphics overlay and mouse connected to the computer can be used to align successive sections via the stepping motors, as well as to trace the outlines of a profile, or of a microtubule.

scholarly journals High-resolution structure of RNase P protein from Thermotoga maritima

2003 ◽  
Vol 100 (13) ◽  
pp. 7497-7502 ◽  
Author(s):  
A. V. Kazantsev ◽  
A. A. Krivenko ◽  
D. J. Harrington ◽  
R. J. Carter ◽  
S. R. Holbrook ◽  
...  
Keyword(s):  
High Resolution ◽  
Thermotoga Maritima ◽  
Rnase P ◽  
P Protein ◽  
High Resolution Structure ◽  
Resolution Structure

scholarly journals A High-Resolution Crystal Structure of a Psychrohalophilic α–Carbonic Anhydrase from Photobacterium profundum Reveals a Unique Dimer Interface

PLoS ONE ◽  
2016 ◽  
Vol 11 (12) ◽  
pp. e0168022 ◽  
Author(s):  
Vijayakumar Somalinga ◽  
Greg Buhrman ◽  
Ashikha Arun ◽  
Robert B. Rose ◽  
Amy M. Grunden
Keyword(s):  
Crystal Structure ◽  
High Resolution ◽  
Carbonic Anhydrase ◽  
Dimer Interface ◽  
Photobacterium Profundum

scholarly journals Asymmetry in catalysis by Thermotoga maritima membrane-bound pyrophosphatase demonstrated by a non-phosphorous allosteric inhibitor

2018 ◽  
Author(s):  
Keni Vidilaseris ◽  
Alexandros Kiriazis ◽  
Ainoleena Turku ◽  
Ayman Khattab ◽  
Niklas G. Johansson ◽  
...  
Keyword(s):  
Drug Target ◽  
Thermotoga Maritima ◽  
Thermophilic Bacterium ◽  
Integral Membrane Proteins ◽  
Sodium Ions ◽  
Exit Channel ◽  
Homologous Proteins ◽  
Inhibitor Binding ◽  
Allosteric Inhibitor ◽  
Membrane Bound

AbstractMembrane-bound pyrophosphatases are homodimeric integral membrane proteins that hydrolyse pyrophosphate into orthophosphates, coupled to the active transport of protons or sodium ions across membranes. They are important in the life cycle of bacteria, archaea, plants, and protist parasites, but no homologous proteins exist in vertebrates, making them a promising drug target. Here, we report the first non-phosphorous allosteric inhibitor (Ki of 1.8 ± 0.3 μM) of the thermophilic bacterium Thermotoga maritima membrane-bound pyrophosphatase and its bound structure at 3.7 Å resolution together with the substrate analogue imidodiphosphate. The unit cell contains two protein homodimers, each binding a single inhibitor dimer near the exit channel, creating a hydrophobic clamp that inhibits the movement of β-strand 1–2 during pumping, and thus preventing the hydrophobic gate from opening. This asymmetry of inhibitor binding with respect to each homodimer provide the first clear demonstration of asymmetry in the catalytic cycle of membrane-bound pyrophosphatases.

scholarly journals Structural insights into archeal-type SAHase from Thermotoga maritima

2014 ◽  
Vol 70 (a1) ◽  
pp. C439-C439
Author(s):  
Patrycja Olszynska ◽  
Monika Imierska ◽  
Justyna Czyrko ◽  
Krzysztof Brzezinski
Keyword(s):  
Thermotoga Maritima ◽  
Enzymatic Reaction ◽  
Thermophilic Bacterium ◽  
Diffusion Method ◽  
Monoclinic Space Group ◽  
X Rays ◽  
Asymmetric Unit ◽  
Cofactor Binding ◽  
Open Conformation ◽  
Structural Insights

S-adenosyl-L-homocysteine hydrolase (SAHase) catalyzes the reversible breakdown of S-adenosyl-L-homocysteine (SAH) to adenosine (Ado) and homocysteine (Hcy). SAH is formed in methylation reactions that utilize S-adenosyl-L-methionine (SAM) as a methyl donor. By removing the SAH byproduct, SAHase serves as a regulator of SAM-dependent biological methylation reactions.[1] Thermotoga maritima is a thermophilic bacterium, but its genome carries a number of archeal genes as a consequence of massive gene transfers related to adaptation to the high-temperature environment.[2] sahh is one of many genes of archeal origin found in T. maritima. Crystals of recombinant SAHase from T. maritima in complex with adenosine were obtained by the hanging drop vapor diffusion method. The crystals are monoclinic, space group C2, with a = 120.4, b =105.5, c = 85.5 Å, β=108.80and diffract X-rays to 1.80 Å. The crystal contains two protein molecules in the asymmetric unit. The enzyme is active as a homotetramer with a molecular weight of about 180 kDa. The crystal contains two protomers in the asymmetric unit, which exist in both, open and closed conformations. The complete tetrameric enzyme molecule is generated in the crystal lattice through the operation of the crystallographic twofold axis. In contrast to other SAHase structures, only two subunits contain a tightly bound NAD+ cofactor, however their closed conformations exclude the possibility of substrate binding. The other two subunits are in open conformation and bind adenosine molecule in the cofactor binding site. Herein, lack of the cofactor molecule excludes the possibility of an enzymatic reaction. In contrast to other SAHases, the C-terminal domain from adjacent protomer does not participate in the binding of the NAD+. Results presented here indicate a different structural organization of archeal type SAHases.

scholarly journals Tunable allosteric library of caspase-3 identifies coupling between conserved water molecules and conformational selection

2016 ◽  
Vol 113 (41) ◽  
pp. E6080-E6088 ◽  
Author(s):  
Joseph J. Maciag ◽  
Sarah H. Mackenzie ◽  
Matthew B. Tucker ◽  
Joshua L. Schipper ◽  
Paul Swartz ◽  
...  
Keyword(s):  
High Resolution ◽  
Posttranslational Modifications ◽  
Caspase 3 ◽  
High Energy ◽  
Water Molecules ◽  
Active Conformation ◽  
Allosteric Site ◽  
Dimer Interface ◽  
Conformational Selection ◽  
Conserved Water Molecules

The native ensemble of caspases is described globally by a complex energy landscape where the binding of substrate selects for the active conformation, whereas targeting an allosteric site in the dimer interface selects an inactive conformation that contains disordered active-site loops. Mutations and posttranslational modifications stabilize high-energy inactive conformations, with mostly formed, but distorted, active sites. To examine the interconversion of active and inactive states in the ensemble, we used detection of related solvent positions to analyze 4,995 waters in 15 high-resolution (<2.0 Å) structures of wild-type caspase-3, resulting in 450 clusters with the most highly conserved set containing 145 water molecules. The data show that regions of the protein that contact the conserved waters also correspond to sites of posttranslational modifications, suggesting that the conserved waters are an integral part of allosteric mechanisms. To test this hypothesis, we created a library of 19 caspase-3 variants through saturation mutagenesis in a single position of the allosteric site of the dimer interface, and we show that the enzyme activity varies by more than four orders of magnitude. Altogether, our database consists of 37 high-resolution structures of caspase-3 variants, and we demonstrate that the decrease in activity correlates with a loss of conserved water molecules. The data show that the activity of caspase-3 can be fine-tuned through globally desolvating the active conformation within the native ensemble, providing a mechanism for cells to repartition the ensemble and thus fine-tune activity through conformational selection.

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