The crystal structure of the heme d
                  1 biosynthesis-associated small c-type cytochrome NirC reveals mixed oligomeric states in crystallo

Monoheme c-type cytochromes are important electron transporters in all domains of life. They possess a common fold hallmarked by three α-helices that surround a covalently attached heme. An intriguing feature of many monoheme c-type cytochromes is their capacity to form oligomers by exchanging at least one of their α-helices, which is often referred to as 3D domain swapping. Here, the crystal structure of NirC, a c-type cytochrome co-encoded with other proteins involved in nitrite reduction by the opportunistic pathogen Pseudomonas aeruginosa, has been determined. The crystals diffracted anisotropically to a maximum resolution of 2.12 Å (spherical resolution of 2.83 Å) and initial phases were obtained by Fe-SAD phasing, revealing the presence of 11 NirC chains in the asymmetric unit. Surprisingly, these protomers arrange into one monomer and two different types of 3D domain-swapped dimers, one of which shows pronounced asymmetry. While the simultaneous observation of monomers and dimers probably reflects the interplay between the high protein concentration required for crystallization and the structural plasticity of monoheme c-type cytochromes, the identification of conserved structural motifs in the monomer together with a comparison with similar proteins may offer new leads to unravel the unknown function of NirC.

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The crystal structure of heme d1 biosynthesis-associated small c-type cytochrome NirC reveals mixed oligomeric states in crystallo

10.1101/2020.02.21.959056 ◽

2020 ◽

Author(s):

Thomas Klünemann ◽

Steffi Henke ◽

Wulf Blankenfeldt

Keyword(s):

Crystal Structure ◽

Pseudomonas Aeruginosa ◽

Opportunistic Pathogen ◽

Nitrite Reduction ◽

Asymmetric Unit ◽

Maximum Resolution ◽

High Protein Concentration ◽

Domains Of Life ◽

Sad Phasing ◽

Heme D1

AbstractMonoheme c-type cytochromes are important electron transporters in all domains of life. They possess a common fold hallmarked by three α-helices that surround a covalently attached heme. An intriguing feature of many monoheme c-type cytochromes is their capacity to form oligomers by exchanging at least one of their α-helices, which is often referred to as 3D domain swapping. Here, we have determined the crystal structure of NirC, a c-type cytochrome co-encoded with other proteins involved in nitrite reduction by the opportunistic pathogen Pseudomonas aeruginosa. Crystals diffracted anisotropically to a maximum resolution of 2.12 Å (spherical resolution 2.83 Å) and initial phases were obtained by Fe-SAD phasing, revealing the presence of eleven NirC chains in the asymmetric unit. Surprisingly, these protomers arrange into one monomer and two different types of 3D-domain-swapped dimers, one showing pronounced asymmetry. While the simultaneous observation of monomers and dimers probably reflects the interplay between high protein concentration required for crystallization and the structural plasticity of monoheme c-type cytochromes, the identification of conserved structural motifs in the monomer together with a comparison to similar proteins may offer new leads to unravel the unknown function of NirC.SynopsisThe crystal structure of the c-type cytochrome NirC from Pseudomonas aeruginosa has been determined and reveals the simultaneous presence of monomers and 3D-domain-swapped dimers in the same asymmetric unit.

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Crystal structure images of large gold clusters and growing crystals by HRTEM

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010011218x ◽

1984 ◽

Vol 42 ◽

pp. 428-429

Author(s):

L.R. Wallenberg ◽

J.-O. Bovin ◽

G. Schmid

Keyword(s):

Crystal Structure ◽

Nucleation And Growth ◽

Close Packing ◽

Gold Clusters ◽

Molecular Weight Determination ◽

Growth Mechanisms ◽

Starting Point ◽

Different Types ◽

Nucleation And Growth Mechanisms ◽

Points Of View

Metallic clusters are interesting from various points of view, e.g. as a mean of spreading expensive catalysts on a support, or following heterogeneous and homogeneous catalytic events. It is also possible to study nucleation and growth mechanisms for crystals with the cluster as known starting point.Gold-clusters containing 55 atoms were manufactured by reducing (C6H5)3PAuCl with B2H6 in benzene. The chemical composition was found to be Au9.2[P(C6H5)3]2Cl. Molecular-weight determination by means of an ultracentrifuge gave the formula Au55[P(C6H5)3]Cl6 A model was proposed from Mössbauer spectra by Schmid et al. with cubic close-packing of the 55 gold atoms in a cubeoctahedron as shown in Fig 1. The cluster is almost completely isolated from the surroundings by the twelve triphenylphosphane groups situated in each corner, and the chlorine atoms on the centre of the 3x3 square surfaces. This gives four groups of gold atoms, depending on the different types of surrounding.

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Engineering nucleosomes for generating diverse chromatin assemblies

Nucleic Acids Research ◽

10.1093/nar/gkab070 ◽

2021 ◽

Author(s):

Zenita Adhireksan ◽

Deepti Sharma ◽

Phoi Leng Lee ◽

Qiuye Bao ◽

Sivaraman Padavattan ◽

...

Keyword(s):

Dynamic Properties ◽

Dna Nanostructures ◽

Macromolecular Assemblies ◽

Maximum Resolution ◽

Different Types ◽

Chromatin Structural ◽

Dna Fragment

Abstract Structural characterization of chromatin is challenging due to conformational and compositional heterogeneity in vivo and dynamic properties that limit achievable resolution in vitro. Although the maximum resolution for solving structures of large macromolecular assemblies by electron microscopy has recently undergone profound increases, X-ray crystallographic approaches may still offer advantages for certain systems. One such system is compact chromatin, wherein the crystalline state recapitulates the crowded molecular environment within the nucleus. Here we show that nucleosomal constructs with cohesive-ended DNA can be designed that assemble into different types of circular configurations or continuous fibers extending throughout crystals. We demonstrate the utility of the method for characterizing nucleosome compaction and linker histone binding at near-atomic resolution but also advance its application for tackling further problems in chromatin structural biology and for generating novel types of DNA nanostructures. We provide a library of cohesive-ended DNA fragment expression constructs and a strategy for engineering DNA-based nanomaterials with a seemingly vast potential variety of architectures and histone chemistries.

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The Native Monomer of Bacillus Pumilus Ribonuclease Does Not Exist Extracellularly

BioMed Research International ◽

10.1155/2018/4837623 ◽

2018 ◽

Vol 2018 ◽

pp. 1-7 ◽

Cited By ~ 7

Author(s):

Olga Ilinskaya ◽

Vera Ulyanova ◽

Irina Lisevich ◽

Elena Dudkina ◽

Nataliya Zakharchenko ◽

...

Keyword(s):

Conformational Changes ◽

Protein Concentration ◽

Bacillus Pumilus ◽

Polyacrylamide Gel Electrophoresis ◽

Culture Fluid ◽

Size Exclusion ◽

High Protein Concentration ◽

Exclusion Chromatography ◽

Native Polyacrylamide Gel Electrophoresis ◽

Hypochromic Effect

Supported by crystallography studies, secreted ribonuclease of Bacillus pumilus (binase) has long been considered to be monomeric in form. Recent evidence obtained using native polyacrylamide gel electrophoresis and size-exclusion chromatography suggests that binase is in fact dimeric. To eliminate ambiguity and contradictions in the data we have measured conformational changes, hypochromic effect, and hydrodynamic radius of binase. The immutability of binase secondary structure upon transition from low to high protein concentration was registered, suggesting the binase dimerization immediately after translocation through the cell membrane and leading to detection of binase dimers only in the culture fluid regardless of ribonuclease concentration. Our results made it necessary to take a fresh look at the binase stability and cytotoxicity towards virus-infected or tumor cells.

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Crystal structure of Pistol, a class of self-cleaving ribozyme

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1611191114 ◽

2017 ◽

Vol 114 (5) ◽

pp. 1021-1026 ◽

Cited By ~ 32

Author(s):

Laura A. Nguyen ◽

Jimin Wang ◽

Thomas A. Steitz

Keyword(s):

Crystal Structure ◽

Rna Structure ◽

Tertiary Structure ◽

Genomic Analysis ◽

Comparative Genomic ◽

Nonbridging Oxygen ◽

Close Proximity ◽

Domains Of Life ◽

Guanine Base ◽

A Minor

Small self-cleaving ribozymes have been discovered in all evolutionary domains of life. They can catalyze site-specific RNA cleavage, and as a result, they have relevance in gene regulation. Comparative genomic analysis has led to the discovery of a new class of small self-cleaving ribozymes named Pistol. We report the crystal structure of Pistol at 2.97-Å resolution. Our results suggest that the Pistol ribozyme self-cleavage mechanism likely uses a guanine base in the active site pocket to carry out the phosphoester transfer reaction. The guanine G40 is in close proximity to serve as the general base for activating the nucleophile by deprotonating the 2′-hydroxyl to initiate the reaction (phosphoester transfer). Furthermore, G40 can also establish hydrogen bonding interactions with the nonbridging oxygen of the scissile phosphate. The proximity of G32 to the O5′ leaving group suggests that G32 may putatively serve as the general acid. The RNA structure of Pistol also contains A-minor interactions, which seem to be important to maintain its tertiary structure and compact fold. Our findings expand the repertoire of ribozyme structures and highlight the conserved evolutionary mechanism used by ribozymes for catalysis.

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Challenges in the development of high protein concentration formulations

Journal of Pharmaceutical Sciences ◽

10.1002/jps.20079 ◽

2004 ◽

Vol 93 (6) ◽

pp. 1390-1402 ◽

Cited By ~ 565

Author(s):

Steven J. Shire ◽

Zahra Shahrokh ◽

Jun Liu

Keyword(s):

Protein Concentration ◽

High Protein ◽

High Protein Concentration

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AAC Richard Soybean

Canadian Journal of Plant Science ◽

10.1139/cjps-2021-0261 ◽

2022 ◽

Author(s):

Kangfu Yu ◽

Lorna Woodrow ◽

M. Chun Shi

Keyword(s):

Glycine Max ◽

Research And Development ◽

Soybean Cyst Nematode ◽

Protein Concentration ◽

Heterodera Glycines ◽

Cyst Nematode ◽

Processing Quality ◽

Food Grade ◽

High Protein Concentration ◽

Glycine Max L

AAC Richard is a food grade soybean [Glycine max (L.) Merr] cultivar with yellow hilum, high protein concentration, and good processing quality for foreign and domestic soymilk, tofu, and miso markets. It has resistance to SCN (soybean cyst nematode) (Heterodera Glycines Ichinohe). AAC Richard was developed at the Agriculture and Agri-Food Canada (AAFC) Harrow Research and Development Centre (Harrow-RDC), Harrow, Ontario and is adapted to areas of southwest Ontario with 3100 or more crop heat units and has a relative maturity of 2.3 (MG 2.3).

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