Structural basis of the catalytic role of Glu301 inAnabaena PCC 7119 ferredoxin-NADP+ reductase revealed by x-ray crystallography

ABSTRACT The psychrophilic alkaline metalloprotease (PAP) produced by a Pseudomonas bacterium isolated in Antarctica belongs to the clan of metzincins, for which a zinc ion is essential for catalytic activity. Binding studies in the crystalline state have been performed by X-ray crystallography in order to improve the understanding of the role of the zinc and calcium ions bound to this protease. Cocrystallization and soaking experiments with EDTA in a concentration range from 1 to 85 mM have resulted in five three-dimensional structures with a distinct number of metal ions occupying the ion-binding sites. Evolution of the structural changes observed in the vicinity of each cation-binding site has been studied as a function of the concentration of EDTA, as well as of time, in the presence of the chelator. Among others, we have found that the catalytic zinc ion was the first ion to be chelated, ahead of a weakly bound calcium ion (Ca 700) exclusive to the psychrophilic enzyme. Upon removal of the catalytic zinc ion, the side chains of the active-site residues His-173, His-179 and Tyr-209 shifted ∼4, 1.0, and 1.6 Å, respectively. Our studies confirm and also explain the sensitivity of PAP toward moderate EDTA concentrations and propose distinct roles for the calcium ions. A new crystal form of native PAP validates our previous predictions regarding the adaptation of this enzyme to cold environments as well as the proteolytic domain calcium ion being exclusive for PAP independent of crystallization conditions.

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A comparative study of 2,4-quinazolinediones by X-ray crystallography. The role of the OMe group on the hydrogen bonding motifs formation

Main Group Chemistry ◽

10.3233/mgc-120078 ◽

2012 ◽

Vol 11 (4) ◽

pp. 259-265

Author(s):

Leticia Guerrero ◽

Ruben Montalvo ◽

Ignacio A. Rivero ◽

Victor Barba

Keyword(s):

Hydrogen Bonding ◽

Comparative Study ◽

X Ray ◽

X Ray Crystallography

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Role of Computational Methods in Going beyond X-ray Crystallography to Explore Protein Structure and Dynamics

International Journal of Molecular Sciences ◽

10.3390/ijms19113401 ◽

2018 ◽

Vol 19 (11) ◽

pp. 3401 ◽

Cited By ~ 16

Author(s):

Ashutosh Srivastava ◽

Tetsuro Nagai ◽

Arpita Srivastava ◽

Osamu Miyashita ◽

Florence Tama

Keyword(s):

Protein Dynamics ◽

Computational Methods ◽

Protein Structures ◽

Three Dimensional ◽

Dimensional Structure ◽

X Ray ◽

X Ray Crystallography ◽

Insight Into

Protein structural biology came a long way since the determination of the first three-dimensional structure of myoglobin about six decades ago. Across this period, X-ray crystallography was the most important experimental method for gaining atomic-resolution insight into protein structures. However, as the role of dynamics gained importance in the function of proteins, the limitations of X-ray crystallography in not being able to capture dynamics came to the forefront. Computational methods proved to be immensely successful in understanding protein dynamics in solution, and they continue to improve in terms of both the scale and the types of systems that can be studied. In this review, we briefly discuss the limitations of X-ray crystallography in studying protein dynamics, and then provide an overview of different computational methods that are instrumental in understanding the dynamics of proteins and biomacromolecular complexes.

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Stereochemical role of lone pairs in main-group elements. Part 3. Structure and bonding in trichloro(1,4,7,10,13-pentaoxacyclopentadecane)antimony(III) studied by means of X-ray crystallography at 120 K

Journal of the Chemical Society Dalton Transactions ◽

10.1039/dt9870000427 ◽

1987 ◽

pp. 427 ◽

Cited By ~ 22

Author(s):

Edward Hough ◽

David G. Nicholson ◽

Asha K. Vasudevan

Keyword(s):

Main Group ◽

Main Group Elements ◽

X Ray ◽

X Ray Crystallography ◽

Lone Pairs ◽

Structure And Bonding

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Structural basis for the binding of SNAREs to the multisubunit tethering complex Dsl1

10.1101/2020.04.07.029496 ◽

2020 ◽

Author(s):

Sophie M. Travis ◽

Kevin DAmico ◽

I-Mei Yu ◽

Safraz Hamid ◽

Gabriel Ramirez-Arellano ◽

...

Keyword(s):

Membrane Trafficking ◽

Relative Orientation ◽

Snare Complex ◽

Structural Basis ◽

Binding Modes ◽

Range Interaction ◽

X Ray ◽

X Ray Crystallography ◽

Long Range Interaction ◽

Target Membrane

AbstractMultisubunit tethering complexes (MTCs) are large (250 to >750 kDa), conserved macromolecular machines that are essential for SNARE-mediated membrane fusion in all eukaryotes. MTCs are thought to function as organizers of membrane trafficking, mediating the initial, long-range interaction between a vesicle and its target membrane and promoting the formation of membrane-bridging SNARE complexes. Previously, we reported the structure of the Dsl1 complex, the simplest known MTC, which is essential for COPI-mediated transport from the Golgi to the endoplasmic reticulum (ER). This structure suggested how the Dsl1 complex might function to tether a vesicle to its target membrane by binding at one end to the COPI coat and at the other end to ER SNAREs. Here, we use x-ray crystallography to investigate these Dsl1-SNARE interactions in greater detail. The Dsl1 complex comprises three subunits that together form a two-legged structure with a central hinge. Our results show that distal regions of each leg bind N-terminal Habc domains of the ER SNAREs Sec20 (a Qb-SNARE) and Use1 (a Qc-SNARE). The observed binding modes appear to anchor the Dsl1 complex to the ER target membrane while simultaneously ensuring that both SNAREs are in open conformations with their SNARE motifs available for assembly. The proximity of the two SNARE motifs, and therefore their ability to enter the same SNARE complex, depends on the relative orientation of the two Dsl1 legs.

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Mechanochemical Synthesis and Structure of the Tetrahydrate and Mesoporous Anhydrous Metforminium(2+)-N,N’-1,4-Phenylenedioxalamic Acid Cocrystal Salt: The Role of Hydrogen Bonding and N→π* Charge Assisted Interactions

10.20944/preprints202009.0660.v1 ◽

2020 ◽

Author(s):

Sayuri Chong-Canto ◽

Efrén V. García-Báez ◽

Francisco J. Martínez-Martínez ◽

Ángel Ramos-Organillo ◽

Itzia I. Padilla-Martínez

Keyword(s):

Hydrogen Bonding ◽

Mechanochemical Synthesis ◽

Water Molecules ◽

X Ray ◽

X Ray Crystallography ◽

Adsorption Desorption ◽

First Time ◽

Selective Formation ◽

Crystallization From Solution

A new cocrystal salt of metformin, an antidiabetic drug, and N,N’-(1,4-phenylene)dioxalamic acid, was synthesized by mechanochemical synthesis, purified by crystallization from solution and characterized by single X-ray crystallography. The structure revealed a salt-type cocrystal composed of one dicationic metformin unit, two monoanionic units of the acid and four water molecules namely H2Mf(HpOXA)2∙4H2O. X-ray powder, IR, 13C-CPMAS, thermal and BET adsorption-desorption analyses were performed to elucidate the structure of the molecular and supramolecurar structure of the anhydrous microcrystalline mesoporous solid H2Mf(HpOXA)2. The results suggest that their structures, conformation and hydrogen bonding schemes are very similar between them. To the best of our knowledge, the selective formation of the monoanion HpOXA⁻, as well as its structure in the solid, is herein reported for the first time. Regular O(-)∙∙∙C(), O(-)∙∙∙N+ and bifacial O(-)∙∙∙C()∙∙∙O(-) of n→* charge-assisted interactions are herein described in H2MfA cocrystal salts which could be responsible of the interactions of metformin in biologic systems. The results, support the participation of n→* charge-assisted interactions independently, and not just as a short contact imposed by the geometric constraint due to the hydrogen bonding patterns.

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Examination of the Catalytic Role of the Axial Cystine Ligand in the Co-Type Nitrile Hydratase from Pseudonocardia thermophila JCM 3095

Catalysts ◽

10.3390/catal11111381 ◽

2021 ◽

Vol 11 (11) ◽

pp. 1381

Author(s):

Irene R. A. M. Ogutu ◽

Martin St. Maurice ◽

Brian Bennett ◽

Richard C. Holz

Keyword(s):

Nitrile Hydratase ◽

Bond Formation ◽

Disulfide Bond Formation ◽

Protein Species ◽

Wild Type ◽

Cobalt Ions ◽

X Ray ◽

Full Complement ◽

Catalytic Role

The strictly conserved αSer162 residue in the Co-type nitrile hydratase from Pseudonocardia thermophila JCM 3095 (PtNHase), which forms a hydrogen bond to the axial αCys108-S atom, was mutated into an Ala residue. The αSer162Ala yielded two different protein species: one was the apoform (αSerA) that exhibited no observable activity, and the second (αSerB) contained its full complement of cobalt ions and was active with a kcat value of 63 ± 3 s−1 towards acrylonitrile at pH 7.5. The X-ray crystal structure of αSerA was determined at 1.85 Å resolution and contained no detectable cobalt per α2β2 heterotetramer. The axial αCys108 ligand itself was also mutated into Ser, Met, and His ligands. All three of these αCys108 mutant enzymes contained only half of the cobalt complement of wild-type PtNHase, but were able to hydrate acrylonitrile with kcat values of 120 ± 6, 29 ± 3, and 14 ± 1 s−1 for the αCys108His, Ser, and Met mutant enzymes, respectively. As all three of these mutant enzymes are catalytically competent, these data provide the first experimental evidence that transient disulfide bond formation is not catalytically essential for NHases.

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A Chemical Probe Based on the PreQ1 Metabolite Enables Transcriptome-wide Mapping of Binding Sites

10.21203/rs.3.rs-322220/v1 ◽

2021 ◽

Author(s):

Sumirtha Balaratnam ◽

Curran Rhodes ◽

Desta Bume ◽

Colleen Connelly ◽

Christopher Lai ◽

...

Keyword(s):

Small Molecule ◽

Binding Sites ◽

X Ray ◽

X Ray Crystallography ◽

Rna Targets ◽

Active Probe ◽

Selective Interaction ◽

Molecule Interactions ◽

Competition Assays

Abstract The role of metabolite-responsive riboswitches in regulating gene expression in bacteria is well known and makes them useful systems for the study of RNA-small molecule interactions. Here, we study the PreQ1 riboswitch system, assessing sixteen diverse PreQ1-derived probes for their ability to selectively modify the riboswitch aptamer covalently. For the most active probe, a diazirine-based photocrosslinker, X-ray crystallography and gel-based competition assays demonstrated the mode of binding of the ligand to the aptamer, and functional assays demonstrated that the probe retains activity against the full riboswitch. Transcriptome-wide mapping using Chem-CLIP revealed a highly selective interaction between the bacterial aptamer and the small molecule. In addition, a small number of RNA targets in endogenous human transcripts were found to bind specifically to PreQ1, providing evidence for candidate PreQ1 aptamers in human RNA. This work demonstrates a stark influence of linker chemistry and structure on the ability of molecules to crosslink RNA, reveals that the PreQ1 aptamer/ligand pair are broadly useful for chemical biology applications, and provides insights into how PreQ1 interacts with human RNAs.

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