Novel T9 loop conformation of filamenting temperature-sensitive mutant Z from Mycobacterium tuberculosis
As of 2017, tuberculosis had infected 1.7 billion people (23% of the population of the world) and caused ten million deaths. Mycobacterium tuberculosis (Mtb) is quickly evolving, and new strains are classified as multidrug resistant. Thus, the identification of novel druggable targets is essential to combat the proliferation of these drug-resistant strains. Filamenting temperature-sensitive mutant Z (FtsZ) is a key protein involved in cytokinesis, an important process for Mtb proliferation and viability. FtsZ is required for bacterial cell division because it polymerizes into a structure called the Z-ring, which recruits a...
Source: Acta Crystallographica Section F - April 23, 2019 Category: Biochemistry Authors: Lazo, E.O. Jakoncic, J. RoyChowdhury, S. Awasthi, D. Ojima, I. Tags: tuberculosis Mycobacterium tuberculosis FtsZ T9 loop filamenting temperature-sensitive mutant Z research communications Source Type: research

Structural basis for oligomerization of the prokaryotic peptide transporter PepTSo2
This study provides valuable insights into the oligomerization mechanism of this MFS-type transporter, which will further pave the way for understanding other oligomeric membrane proteins. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 23, 2019 Category: Biochemistry Authors: Nagamura, R. Fukuda, M. Kawamoto, A. Matoba, K. Dohmae, N. Ishitani, R. Takagi, J. Nureki, O. Tags: peptide transporter major facilitator superfamily lipidic cubic phase single-particle cryo-electron microscopy membrane transporter oligomerization cryo-EM X-ray crystallography research communications Source Type: research

Cryo-EM structure of the Ebola virus nucleoprotein – RNA complex
Ebola virus is an emerging virus that is capable of causing a deadly disease in humans. Replication, transcription and packaging of the viral genome are carried out by the viral nucleocapsid. The nucleocapsid is a complex of the viral nucleoprotein, RNA and several other viral proteins. The nucleoprotein forms large, RNA-bound, helical filaments and acts as a scaffold for additional viral proteins. The 3.1   Å resolution single-particle cryo-electron microscopy structure of the nucleoprotein – RNA helical filament presented here resembles previous structures determined at lower resolution, while providing imp...
Source: Acta Crystallographica Section F - April 23, 2019 Category: Biochemistry Authors: Kirchdoerfer, R.N. Saphire, E.O. Ward, A.B. Tags: Ebola virus nucleoprotein NP helix cryo-electron microscopy research communications Source Type: research

Noncovalent structure of SENP1 in complex with SUMO2
SUMOylation is a post-translational modification in which a small ubiquitin-like molecule (SUMO) is appended to substrate proteins and is known to influence myriads of biological processes. A delicate interplay between several families of SUMOylation proteins and their substrates ensures the proper level of SUMOylation required for normal cell function. Among the SUMO proteins, SUMO2 is known to form mono-SUMOylated proteins and engage in poly-SUMO chain formation, while sentrin-specific protease 1 (SENP1) is a key enzyme in regulating both events. Determination of the SENP1 – SUMO2 interaction is therefore necessary...
Source: Acta Crystallographica Section F - April 23, 2019 Category: Biochemistry Authors: Ambaye, N.D. Tags: SENP1 SUMO2 protease SUMOylation X-ray diffraction cancer research communications Source Type: research

Crystal structure of the cold-adapted haloalkane dehalogenase DpcA from Psychrobacter cryohalolentis K5
Haloalkane dehalogenases (HLDs) convert halogenated aliphatic pollutants to less toxic compounds by a hydrolytic mechanism. Owing to their broad substrate specificity and high enantioselectivity, haloalkane dehalogenases can function as biosensors to detect toxic compounds in the environment or can be used for the production of optically pure compounds. Here, the structural analysis of the haloalkane dehalogenase DpcA isolated from the psychrophilic bacterium Psychrobacter cryohalolentis K5 is presented at the atomic resolution of 1.05   Å . This enzyme exhibits a low temperature optimum, making it attractive for e...
Source: Acta Crystallographica Section F - April 23, 2019 Category: Biochemistry Authors: Tratsiak, K. Prudnikova, T. Drienovska, I. Damborsky, J. Brynda, J. Pachl, P. Kuty, M. Chaloupkova, R. Rezacova, P. Kuta Smatanova, I. Tags: haloalkane dehalogenase α / β -hydrolase X-ray diffraction psychrophiles structural analysis Psychrobacter cryohalolentis research communications Source Type: research

The putative polysaccharide deacetylase Ba0331: cloning, expression, crystallization and structure determination
Ba0331 is a putative polysaccharide deacetylase from Bacillus anthracis, the etiological agent of the disease anthrax, that contributes to adaptation of the bacterium under extreme conditions and to maintenance of the cell shape. In the present study, the crystal structure of Ba0331 was determined at 2.6   Å resolution. The structure consists of two domains: a fibronectin type 3-like (Fn3-like) domain and a NodB catalytic domain. The latter is present in all carbohydrate esterase family 4 enzymes, while a comparative analysis of the Fn3-like domain revealed structural plasticity despite the retention of the conserv...
Source: Acta Crystallographica Section F - April 2, 2019 Category: Biochemistry Authors: Andreou, A. Giastas, P. Arnaouteli, S. Tzanodaskalaki, M. Tzartos, S.J. Bethanis, K. Bouriotis, V. Eliopoulos, E.E. Tags: Bacillus anthracis Ba0331 X-ray crystal structure metalloenzymes CE4 family lipoprotein polysaccharide deacetylase research communications Source Type: research

Igni18, a novel metallo-hydrolase from the hyperthermophilic archaeon Ignicoccus hospitalis KIN4/I: cloning, expression, purification and X-ray analysis
The hyperthermophilic crenarchaeon Ignicoccus hospitalis KIN4/I possesses at least 35 putative genes encoding enzymes that belong to the α / β -hydrolase superfamily. One of those genes, the metallo-hydrolase-encoding igni18, was cloned and heterologously expressed in Pichia pastoris. The enzyme produced was purified in its catalytically active form. The recombinant enzyme was successfully crystallized and the crystal diffracted to a resolution of 2.3   Å . The crystal belonged to space group R32, with unit-cell parameters a = b = 67.42, c   =   253.77   Å , α = β = 90.0, &ga...
Source: Acta Crystallographica Section F - April 1, 2019 Category: Biochemistry Authors: Kobus, S. Perez-Garcia, P. Hoeppner, A. Holzscheck, N. Kovacic, F. Streit, W.R. Jaeger, K.-E. Chow, J. Smits, S.H.J. Tags: Ignicoccus hospitalis α / β -hydrolases metallo- -lactamases research communications Source Type: research

Functional and structural characterization of IdnL7, an adenylation enzyme involved in incednine biosynthesis
Adenylation enzymes play an important role in the selective incorporation of the cognate carboxylate substrates in natural product biosynthesis. Here, the biochemical and structural characterization of the adenylation enzyme IdnL7, which is involved in the biosynthesis of the macrolactam polyketide antibiotic incednine, is reported. Biochemical analysis showed that IdnL7 selects and activates several small amino acids. The structure of IdnL7 in complex with an l-alanyl-adenylate intermediate mimic, 5 ′ -O-[N-(l-alanyl)sulfamoyl]adenosine, was determined at 2.1   Å resolution. The structure of IdnL7 explains t...
Source: Acta Crystallographica Section F - April 1, 2019 Category: Biochemistry Authors: Cie ś lak, J. Miyanaga, A. Takaishi, M. Kudo, F. Eguchi, T. Tags: macrolactam antibiotics polyketides crystal structure adenylation enzyme natural product biosynthesis research communications Source Type: research

Structure of a critical metabolic enzyme: S-adenosylmethionine synthetase from Cryptosporidium parvum
S-Adenosyl-l-methionine (AdoMet), the primary methyl donor in most biological methylation reactions, is produced from ATP and methionine in a multistep reaction catalyzed by AdoMet synthetase. The diversity of group-transfer reactions that involve AdoMet places this compound at a key crossroads in amino-acid, nucleic acid and lipid metabolism, and disruption of its synthesis has adverse consequences for all forms of life. The family of AdoMet synthetases is highly conserved, and structures of this enzyme have been determined from organisms ranging from bacteria to humans. Here, the structure of an AdoMet synthetase from th...
Source: Acta Crystallographica Section F - April 1, 2019 Category: Biochemistry Authors: Ohren, J. Parungao, G.F. Viola, R.E. Tags: enzyme structure S-adenosylmethionine synthetase antiparasitic drugs subunit interface Cryptosporidium parvum research communications Source Type: research

Crystal structure of phosphoribulokinase from Synechococcus sp. strain PCC 6301
Phosphoribulokinase (PRK) catalyses the ATP-dependent phosphorylation of ribulose 5-phosphate to give ribulose 1,5-bisphosphate. Regulation of this reaction in response to light controls carbon fixation during photosynthesis. Here, the crystal structure of PRK from the cyanobacterium Synechococcus sp. strain PCC 6301 is presented. The enzyme is dimeric and has an α / β -fold with an 18-stranded β -sheet at its core. Interestingly, a disulfide bond is found between Cys40 and the P-loop residue Cys18, revealing the structural basis for the redox inactivation of PRK activity. A second disulfide bond appears to...
Source: Acta Crystallographica Section F - April 1, 2019 Category: Biochemistry Authors: Wilson, R.H. Hayer-Hartl, M. Bracher, A. Tags: phosphoribulokinase Calvin – Benson Bassham cycle photosynthesis dark reaction redox regulation transferases research communications Source Type: research

Pseudomonas aeruginosa esterase PA2949, a bacterial homolog of the human membrane esterase ABHD6: expression, purification and crystallization
This study provides hints on the functional similarity of ABHD6-like proteins in prokaryotes and eukaryotes, and might guide the structural study of these difficult-to-crystallize proteins. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 1, 2019 Category: Biochemistry Authors: Bleffert, F. Granzin, J. Gohlke, H. Batra-Safferling, R. Jaeger, K.-E. Kovacic, F. Tags: Pseudomonas aeruginosa PA2949 esterases ABHD6 endocannabinoid signalling pathway Parkinson's disease Alzheimer's disease research communications Source Type: research

Perdeuteration, large crystal growth and neutron data collection of Leishmania mexicana triose-phosphate isomerase E65Q variant
Triose-phosphate isomerase (TIM) catalyses the interconversion of dihydroxyacetone phosphate and glyceraldehyde 3-phosphate. Two catalytic mechanisms have been proposed based on two reaction-intermediate analogues, 2-phosphoglycolate (2PG) and phosphoglycolohydroxamate (PGH), that have been used as mimics of the cis-enediol(ate) intermediate in several studies of TIM. The protonation states that are critical for the mechanistic interpretation of these structures are generally not visible in the X-ray structures. To resolve these questions, it is necessary to determine the hydrogen positions using neutron crystallography. N...
Source: Acta Crystallographica Section F - April 1, 2019 Category: Biochemistry Authors: Kelp š as, V. Lafumat, B. Blakeley, M.P. Coquelle, N. Oksanen, E. von Wachenfeldt, C. Tags: triose-phosphate isomerase neutron diffraction Laue method large crystals perdeuteration research communications Source Type: research

Crystallization of the human tetraspanin protein CD9
In this study, attempts were made to crystallize human CD9, a representative member of the tetraspanin family, and it was demonstrated that the truncation of a variable region in the second long extracellular loop significantly improved crystal growth. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 1, 2019 Category: Biochemistry Authors: Umeda, R. Nishizawa, T. Nureki, O. Tags: crystallization LCP membrane proteins CD9 tetraspanins research communications Source Type: research

Crystal structure of the putative peptide-binding protein AppA from Clostridium difficile
Peptides play an important signalling role in Bacillus subtilis, where their uptake by one of two ABC-type oligopeptide transporters, Opp and App, is required for efficient sporulation. Homologues of these transporters in Clostridium difficile have been characterized, but their role, and hence that of peptides, in regulating sporulation in this organism is less clear. Here, the oligopeptide-binding receptor proteins for these transporters, CdAppA and CdOppA, have been purified and partially characterized, and the crystal structure of CdAppA has been determined in an open unliganded form. Peptide binding to either protein c...
Source: Acta Crystallographica Section F - March 12, 2019 Category: Biochemistry Authors: Hughes, A. Wilson, S. Dodson, E.J. Turkenburg, J.P. Wilkinson, A.J. Tags: Clostridium difficile peptide transport OppA AppA sporulation research communications Source Type: research

BacMam production and crystal structure of nonglycosylated apo human furin at 1.89   Å resolution
Furin, also called proprotein convertase subtilisin/kexin 3 (PCSK3), is a calcium-dependent serine endoprotease that processes a wide variety of proproteins involved in cell function and homeostasis. Dysregulation of furin has been implicated in numerous disease states, including cancer and fibrosis. Mammalian cell expression of the furin ectodomain typically produces a highly glycosylated, heterogeneous protein, which can make crystallographic studies difficult. Here, the expression and purification of nonglycosylated human furin using the BacMam technology and site-directed mutagenesis of the glycosylation sites is repor...
Source: Acta Crystallographica Section F - March 12, 2019 Category: Biochemistry Authors: Pearce, K.H. Overton, L.K. Gampe, R.T. Barrett, G.B. Taylor, J.D. McKee, D.D. Campobasso, N. Nolte, R.T. Reid, R.A. Tags: furin nonglycosylated apo serine proteinase hydrolase BacMam research communications Source Type: research

Crystal structure of protein tyrosine phosphatase-2 from Cydia pomonella granulovirus
Many viral genomes encode kinase and phosphatase enzymes to manipulate pathways that are controlled by phosphorylation events. The majority of viral phosphatase genes occur in the Baculoviridae and Poxviridae families of large DNA viruses. The corresponding protein sequences belong to four major homology groups, and structures are currently available for only two of these. Here, the first structure from the third group, the protein tyrosine phosphatase-2 (PTP-2) class of viral phosphatases, is described. It is shown that Cydia pomonella granulovirus PTP-2 has the same general fold and active-site architecture as described ...
Source: Acta Crystallographica Section F - March 12, 2019 Category: Biochemistry Authors: Huang, G. Oliver, M.R. Keown, J.R. Goldstone, D.C. Metcalf, P. Tags: Cydia pomonella granulovirus PTP-2 protein tyrosine phosphatase-2 phosphatases research communications Source Type: research

The structure of lipopolysaccharide transport protein B (LptB) from Burkholderia pseudomallei
The thick outer membrane (OM) of Gram-negative bacteria performs an important protective role against hostile environments, supports cell integrity, and contributes to surface adhesion and in some cases also to virulence. A major component of the OM is lipopolysaccharide (LPS), a complex glycolipid attached to a core containing fatty-acyl chains. The assembly and transport of lipid A, the membrane anchor for LPS, to the OM begins when a heteromeric LptB2FG protein complex extracts lipid A from the outer leaflet of the inner membrane. This process requires energy, and upon hydrolysis of ATP one component of the heteromeric ...
Source: Acta Crystallographica Section F - March 12, 2019 Category: Biochemistry Authors: Pankov, G. Dawson, A. Hunter, W.N. Tags: ABC transporters ATPases lipopolysaccharide transport complex protein – protein interactions research communications Source Type: research

The structure of an iron-containing alcohol dehydrogenase from a hyperthermophilic archaeon in two chemical states
An iron-containing alcohol dehydrogenase (FeADH) from the hyperthermophilic archaeon Thermococcus thioreducens was crystallized in unit cells belonging to space groups P21, P212121 and P43212, and the crystal structures were solved at 2.4, 2.1 and 1.9   Å resolution, respectively, by molecular replacement using the FeADH from Thermotoga maritima (Schwarzenbacher et al., 2004) as a model. In the monoclinic and orthorhombic crystals the dehydrogenase (molecular mass 41.5   kDa) existed as a dimer containing a twofold noncrystallographic symmetry axis, which was crystallographic in the tetragonal crystals. In the mo...
Source: Acta Crystallographica Section F - March 12, 2019 Category: Biochemistry Authors: Larson, S.B. Jones, J.A. McPherson, A. Tags: NADP oxidation – reduction thermophile active site dimers alcohol dehydrogenase Thermococcus thioreducens research communications Source Type: research

Substrate-analogue complex structure of Mycobacterium tuberculosis decaprenyl diphosphate synthase
Decaprenyl diphosphate synthase from Mycobacterium tuberculosis (MtDPPS, also known as Rv2361c) catalyzes the consecutive elongation of ω ,E,Z-farnesyl diphosphate (EZ-FPP) by seven isoprene units by forming new cis double bonds. The protein folds into a butterfly-like homodimer like most other cis-type prenyltransferases. The starting allylic substrate EZ-FPP is bound to the S1 site and the homoallylic substrate to be incorporated, isopentenyl diphosphate, is bound to the S2 site. Here, a 1.55   Å resolution structure of MtDPPS in complex with the substrate analogues geranyl S-thiodiphosphate (GSPP) and isop...
Source: Acta Crystallographica Section F - March 12, 2019 Category: Biochemistry Authors: Ko, T.-P. Xiao, X. Guo, R.-T. Huang, J.-W. Liu, W. Chen, C.-C. Tags: Rv2361c cis-prenyltransferase catalytic mechanism thiodiphosphate inhibitor cell-wall biosynthesis Mycobacterium tuberculosis decaprenyl diphosphate synthase research communications Source Type: research

Welcoming Janet Newman with a BLAST on crystallization strategy
(Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - March 1, 2019 Category: Biochemistry Authors: van Raaij, M.J. Tags: editorial crystallization Source Type: research

The structure of CgnJ, a domain of unknown function protein from the crocagin gene cluster
Natural products often contain interesting new chemical entities that are introduced into the structure of a compound by the enzymatic machinery of the producing organism. The recently described crocagins are novel polycyclic peptides which belong to the class of ribosomally synthesized and post-translationally modified peptide natural products. They have been shown to bind to the conserved prokaryotic carbon-storage regulator A in vitro. In efforts to understand crocagin biosynthesis, the putative biosynthetic genes were expressed and purified. Here, the first crystal structure of a protein from the crocagin-biosynthetic ...
Source: Acta Crystallographica Section F - February 21, 2019 Category: Biochemistry Authors: Adam, S. Klein, A. Surup, F. Koehnke, J. Tags: crocagin RiPPs CgnJ domains of unknown function research communications Source Type: research

Crystal structure of the legume lectin-like domain of an ERGIC-53-like protein from Entamoeba histolytica
ERGIC-53-like proteins are type I membrane proteins that belong to the class of intracellular cargo receptors and are known to be indispensable for the intracellular transport of glycoproteins. They are implicated in transporting glycoproteins between the endoplasmic reticulum and the Golgi body. The crystal structure of the legume lectin-like domain of an ERGIC-53-like protein from Entamoeba histolytica has been determined at 2.4   Å resolution. Although the overall structure of the domain resembles those of its mammalian and yeast orthologs (ERGIC-53 and Emp46, respectively), there are significant changes in the ...
Source: Acta Crystallographica Section F - February 21, 2019 Category: Biochemistry Authors: Khan, F. Suguna, K. Tags: ERGIC-53-like proteins legume lectin-like domains Entamoeba histolytica membrane proteins intracellular cargo receptors research communications Source Type: research

Neutron crystallographic study of heterotrimeric glutamine amidotransferase CAB
This study represented a challenge in current neutron diffraction technology. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - February 21, 2019 Category: Biochemistry Authors: Li, L. Adachi, M. Yu, J. Kato, K. Shinoda, A. Ostermann, A. Schrader, T.E. Ose, T. Yao, M. Tags: glutamine amidotransferase CAB GatCAB ammonia-self-sufficient mechanism ammonia channel neutron diffraction research communications Source Type: research

BLASTing away preconceptions in crystallization trials
Crystallization is in many cases a critical step for solving the three-dimensional structure of a protein molecule. Determining which set of chemicals to use in the initial screen is typically agnostic of the protein under investigation; however, crystallization efficiency could potentially be improved if this were not the case. Previous work has assumed that sequence similarity may provide useful information about appropriate crystallization cocktails; however, the authors are not aware of any quantitative verification of this assumption. This research investigates whether, given current information, one can detect any co...
Source: Acta Crystallographica Section F - February 21, 2019 Category: Biochemistry Authors: Abrahams, G.J. Newman, J. Tags: PDB Protein Data Bank REMARK 280 crystallization sequences BLAST research communications Source Type: research

Structure of glycerol dehydrogenase (GldA) from Escherichia coli
Escherichia coli (strain K-12, substrain MG1655) glycerol dehydrogenase (GldA) is required to catalyze the first step in fermentative glycerol metabolism. The protein was expressed and purified to homogeneity using a simple combination of heat-shock and chromatographic methods. The high yield of the protein ( ∼ 250   mg per litre of culture) allows large-scale production for potential industrial applications. Purified GldA exhibited a homogeneous tetrameric state ( ∼ 161   kDa) in solution and relatively high thermostability (Tm = 65.6 ° C). Sitting-drop sparse-matrix screens were used for protein crystalli...
Source: Acta Crystallographica Section F - February 21, 2019 Category: Biochemistry Authors: Zhang, J. Nanjaraj Urs, A.N. Lin, L. Zhou, Y. Hu, Y. Hua, G. Gao, Q. Yuchi, Z. Zhang, Y. Tags: glycerol dehydrogenase thermal stability crystal structure glycerol metabolism research communications Source Type: research

Neutron and X-ray analysis of the Fenna – Matthews – Olson photosynthetic antenna complex from Prosthecochloris aestuarii
The Fenna – Matthews – Olson protein from Prosthecochloris aestuarii (PaFMO) has been crystallized in a new form that is amenable to high-resolution X-ray and neutron analysis. The crystals belonged to space group H3, with unit-cell parameters a = b = 83.64, c = 294.78   Å , and diffracted X-rays to ∼ 1.7   Å resolution at room temperature. Large PaFMO crystals grown to volumes of 0.3 – 0.5   mm3 diffracted neutrons to 2.2   Å resolution on the MaNDi neutron diffractometer at the Spallation Neutron Source. The resolution of the neutron data will allow direct determination of ...
Source: Acta Crystallographica Section F - February 20, 2019 Category: Biochemistry Authors: Lu, X. Selvaraj, B. Ghimire-Rijal, S. Orf, G.S. Meilleur, F. Blankenship, R.E. Cuneo, M.J. Myles, D.A.A. Tags: neutron crystallography photosynthesis hydrogen site energy neutron diffraction Fenna – Matthews Olson protein Prosthecochloris aestuarii research communications Source Type: research

Carbonic anhydrase II in complex with carboxylic acid-based inhibitors
This study examines an important class of alternative, non-sulfonamide-based CA inhibitors and provides insight to facilitate the structure-guided design of CA isoform-specific inhibitors. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - February 20, 2019 Category: Biochemistry Authors: Lomelino, C.L. McKenna, R. Tags: carbonic anhydrase II carboxylic acids structure-guided drug design research communications Source Type: research

Preparation of stable recombinant Osm1 noncovalently bound with flavin adenosine dinucleotide cofactor for structural study
In this study, monomeric and stable recombinant Osm1 was successfully prepared for structural study. During purification, it was realized that the majority of recombinant Osm1 expressed in Escherichia coli lacked the flavin adenosine dinucleotide (FAD) cofactor. However, exogenously introduced FAD could be incorporated into recombinant Osm1, generating stable and homogenous holo Osm1. Moreover, after removing a flexible fragment by limited proteolysis, holo Osm1 formed isotropic crystals that retained catalytic activity. X-ray diffraction data were successfully collected from the Osm1 crystals to a resolution of 1.75   &...
Source: Acta Crystallographica Section F - February 20, 2019 Category: Biochemistry Authors: Kim, S.H. Park, H.H. Tags: soluble fumarate reductase anaerobiosis Osm1 limited proteolysis stabilization crystallization research communications Source Type: research

Crystal structure of the type VI immunity protein Tdi1 (Atu4351) from Agrobacterium tumefaciens
The type VI secretion system (T6SS) is a novel multiprotein needle-like apparatus that is distributed widely in Gram-negative bacteria. Bacteria harboring T6SSs inject various effectors into both eukaryotic and prokaryotic cells for interspecies competition or virulence-related processes. The toxicities of the effectors can be neutralized by their cognate immunity proteins. Tde1 (Atu4350) – Tdi1 (Atu4351) has recently been characterized as a T6SS effector – immunity pair in the soil bacterium Agrobacterium tumefaciens and the neutralization mechanism remains unknown. Here, the crystal structure of the immunity ...
Source: Acta Crystallographica Section F - February 20, 2019 Category: Biochemistry Authors: Shi, L. Gao, Z. Zhang, T. Zhang, H. Dong, Y. Tags: type VI secretion system effector – immunity pair Tdi1 GAD-like domain DUF1851 domain bacterial nanomachines research communications Source Type: research

An assessment of three human methylenetetrahydrofolate dehydrogenase/cyclohydrolase – ligand complexes following further refinement
The enzymes involved in folate metabolism are key drug targets for cell-growth modulation, and accurate crystallographic structures provide templates to be exploited for structure-based ligand design. In this context, three ternary complex structures of human methylenetetrahydrofolate dehydrogenase/cyclohydrolase have been published [Schmidt et al. (2000), Biochemistry, 39, 6325 – 6335] and potentially represent starting points for the development of new antifolate inhibitors. However, an inspection of the models and the deposited data revealed deficiencies and raised questions about the validity of the structures. A...
Source: Acta Crystallographica Section F - February 20, 2019 Category: Biochemistry Authors: Bueno, R. Dawson, A. Hunter, W.N. Tags: methylenetetrahydrofolate dehydrogenase methenyltetrahydrofolate cyclohydrolase antifolates structure-based ligand design structure validation re-refinement research communications Source Type: research

Crystal structure of the aromatic-amino-acid aminotransferase from Streptococcus mutans
In this study, the aromatic-amino-acid aminotransferase from Streptococcus mutans (SmAroAT) was recombinantly expressed in Escherichia coli. An effective purification protocol was established. The recombinant protein was crystallized using the hanging-drop vapor-diffusion method with PEG 3350 as the primary precipitant. The crystal structure of SmAroAT was solved at 2.2   Å resolution by the molecular-replacement method. Structural analysis indicated that the proteins of the aromatic-amino-acid aminotransferase family have conserved structural elements that might play a role in substrate binding. These results may ...
Source: Acta Crystallographica Section F - January 24, 2019 Category: Biochemistry Authors: Cong, X. Li, X. Li, S. Tags: Streptococcus mutans dental caries dental plaque infective endocarditis aromatic-amino-acid aminotransferase crystallization structure determination research communications Source Type: research

Penetration of dyes into protein crystals
Experiments were carried out on 15 different protein crystals with the objective of estimating the rates of penetration of dye molecules into the crystals. The dyes were in the molecular-weight range 250 – 1000   Da and the protein crystals were of dimensions of 0.7   mm or greater. Experiments were also conducted on protein crystals grown between glass cover slips (separation 200   µ m) that restricted the direction of diffusion. The rate of penetration of dyes into protein crystals depends very much on the degree of association between the dye and protein molecules. Dye penetration was not consistent wi...
Source: Acta Crystallographica Section F - January 24, 2019 Category: Biochemistry Authors: McPherson, A. Tags: penetration rates diffusion rates stains microscopy color time lapse research communications Source Type: research

Iterative screen optimization maximizes the efficiency of macromolecular crystallization
Advances in X-ray crystallography have streamlined the process of determining high-resolution three-dimensional macromolecular structures. However, a rate-limiting step in this process continues to be the generation of crystals that are of sufficient size and quality for subsequent diffraction experiments. Here, iterative screen optimization (ISO), a highly automated process in which the precipitant concentrations of each condition in a crystallization screen are modified based on the results of a prior crystallization experiment, is described. After designing a novel high-throughput crystallization screen to take full adv...
Source: Acta Crystallographica Section F - January 24, 2019 Category: Biochemistry Authors: Jones, H.G. Wrapp, D. Gilman, M.S.A. Battles, M.B. Wang, N. Sacerdote, S. Chuang, G.-Y. Kwong, P.D. McLellan, J.S. Tags: automated liquid handling crystallization screening macromolecular crystallography Sweet16 research communications Source Type: research

High-resolution structure of a Y27W mutant of the Dishevelled2 DIX domain
Dishevelled (Dvl) is a positive regulator of the canonical Wnt pathway that downregulates the phosphorylation of β -catenin and its subsequent degradation. Dvl contains an N-terminal DIX domain, which is involved in its homooligomerization and interactions with regulators of the Wnt pathway. The crystal structure of a Y27W mutant of the Dishevelled2 DIX domain (DIX-Y27W) has been determined at 1.64   Å resolution. DIX-Y27W has a compact ubiquitin-like fold and self-associates with neighbouring molecules through β -bridges, resulting in a head-to-tail helical molecular arrangement similar to previously rep...
Source: Acta Crystallographica Section F - January 23, 2019 Category: Biochemistry Authors: Yamanishi, K. Sin, Y. Terawaki, S. Higuchi, Y. Shibata, N. Tags: Wnt signalling pathway Dishevelled DIX domain X-ray structure analysis polymerizing domain research communications Source Type: research

Titration of ionizable groups in proteins using multiple neutron data sets from a single crystal: application to the small GTPase Ras
Neutron protein crystallography (NPC) reveals the three-dimensional structures of proteins, including the positions of H atoms. The technique is particularly suited to elucidate ambiguous catalytic steps in complex biochemical reactions. While NPC uniquely complements biochemical assays and X-ray structural analyses by revealing the protonation states of ionizable groups at and around the active site of enzymes, the technique suffers from a major drawback: large single crystals must be grown to compensate for the relatively low flux of neutron beams. However, in addition to revealing the positions of hydrogens involved in ...
Source: Acta Crystallographica Section F - January 23, 2019 Category: Biochemistry Authors: Knihtila, R. Volmar, A.Y. Meilleur, F. Mattos, C. Tags: in crystallo titration neutron crystallography pKa protonation radiation damage research communications Source Type: research

Crystallization and X-ray analysis of monodisperse human properdin
The 54   kDa protein properdin, also known as factor P (FP), plays a major role in the complement system through the stabilization of the alternative pathway convertases. FP circulates in the blood as cyclic dimers, trimers and tetramers, and this heterogeneity challenges detailed structural insight into the mechanism of convertase stabilization by FP. Here, the generation of an intact FP monomer and a variant monomer with the third thrombospondin repeat liberated is described. Both FP monomers were excised from recombinant full-length FP containing internal cleavage sites for TEV protease. These FP monomers could be cry...
Source: Acta Crystallographica Section F - January 23, 2019 Category: Biochemistry Authors: Pedersen, D.V. Revel, M. Gadeberg, T.A.F. Andersen, G.R. Tags: complement properdin modular proteins crystallization research communications Source Type: research

A revisited version of the apo structure of the ligand-binding domain of the human nuclear receptor retinoic X receptor α
The retinoic X receptor (RXR) plays a crucial role in the superfamily of nuclear receptors (NRs) by acting as an obligatory partner of several nuclear receptors; its role as a transcription factor is thus critical in many signalling pathways, such as metabolism, cell development, differentiation and cellular death. The first published structure of the apo ligand-binding domain (LBD) of RXR α , which is still used as a reference today, contained inaccuracies. In the present work, these inaccuracies were corrected using modern crystallographic tools. The most important correction concerns the presence of a π -bulge ...
Source: Acta Crystallographica Section F - January 23, 2019 Category: Biochemistry Authors: Eberhardt, J. McEwen, A.G. Bourguet, W. Moras, D. Dejaegere, A. Tags: nuclear receptors retinoic acid ligand binding domains retinoic X receptor α research communications Source Type: research

Structure of an Influenza A virus N9 neuraminidase with a tetrabrachion-domain stalk
The influenza neuraminidase (NA) is a homotetramer with head, stalk, transmembrane and cytoplasmic regions. The structure of the NA head with a stalk has never been determined. The NA head from an N9 subtype influenza A virus, A/tern/Australia/G70C/1975 (H1N9), was expressed with an artificial stalk derived from the tetrabrachion (TB) tetramerization domain from Staphylothermus marinus. The NA was successfully crystallized both with and without the TB stalk, and the structures were determined to 2.6 and 2.3   Å resolution, respectively. Comparisons of the two NAs with the native N9 NA structure from egg-grown virus...
Source: Acta Crystallographica Section F - January 23, 2019 Category: Biochemistry Authors: Streltsov, V.A. Schmidt, P.M. McKimm-Breschkin, J.L. Tags: Influenza A virus neuraminidases neuraminidase stalk tetrabrachion tetramerization domain N9 neuraminidase structure research communications Source Type: research

Structures of the antibody 64M-5 Fab and its complex with dT(6 – 4)T indicate induced-fit and high-affinity mechanisms
DNA photoproducts with (6 – 4) pyrimidine – pyrimidone adducts produced by ultraviolet light are mutagenic and carcinogenic. The crystal structures of the anti-(6 – 4) photoproduct antibody 64M-5 Fab and of its complex with dT(6 – 4)T were determined at 2.5 and 2.0   Å resolution, respectively. A comparison between the dT(6 – 4)T-liganded and unliganded structures indicates that the side chain of His93L is greatly rotated and shifted on binding to dT(6 – 4)T, leading to the formation of an electrostatic interaction with the phosphate moiety of dT(6 – 4)T, which shows a rema...
Source: Acta Crystallographica Section F - January 23, 2019 Category: Biochemistry Authors: Yokoyama, H. Mizutani, R. Noguchi, S. Hayashida, N. Tags: antibodies Fabs crystal structure DNA (6 – 4) photoproduct ultraviolet light research communications Source Type: research

Crystal structure of the programmed cell death 5 protein from Sulfolobus solfataricus
Programmed cell death 5 (PDCD5) is a vital signaling protein in the apoptosis pathway in eukaryotes. It is known that there are two dissociated N-terminal regions and a triple-helix core in eukaryotic PDCD5. Structural and functional studies of PDCD5 from hyperthermophilic archaea have been limited to date. Here, the PDCD5 homolog Sso0352 (SsoPDCD5) was identified in Sulfolobus solfataricus, the SsoPDCD5 protein was expressed and crystallized, and the phase was identified by single-wavelength anomalous diffraction. The native SsoPDCD5 crystal belonged to space group C2 and diffracted to 1.49   Å resolution. This is...
Source: Acta Crystallographica Section F - January 23, 2019 Category: Biochemistry Authors: Lin, K.-F. Hsu, J.-Y. Hsieh, D.-L. Tsai, M.-J. Yeh, C.-H. Chen, C.-Y. Tags: Sulfolobus solfataricus DNA-binding protein programmed cell death X-ray crystallography PDCDS research communications Source Type: research

Visualization of biological macromolecules at near-atomic resolution: cryo-electron microscopy comes of age
Structural biology is going through a revolution as a result of transformational advances in the field of cryo-electron microscopy (cryo-EM) driven by the development of direct electron detectors and ultrastable electron microscopes. High-resolution cryo-EM images of isolated biomolecules (single particles) suspended in a thin layer of vitrified buffer are subjected to powerful image-processing algorithms, enabling near-atomic resolution structures to be determined in unprecedented numbers. Prior to these advances, electron crystallography of two-dimensional crystals and helical assemblies of proteins had established the f...
Source: Acta Crystallographica Section F - December 24, 2018 Category: Biochemistry Authors: Mitra, A.K. Tags: cryo-EM single-particle analysis three-dimensional reconstruction phase plates direct detectors topical reviews Source Type: research

Acta Crystallographica Section F – another home for cryo-electron microscopy contributions
(Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - December 24, 2018 Category: Biochemistry Authors: Mitra, A.K. van Raaij, M. Tags: cryo-EM introduction editorial Source Type: research

AtNPR4 from Arabidopsis thaliana: expression, purification, crystallization and crystallographic analysis
In this study, Spodoptera frugiperda (Sf9) cells were used to express full-length AtNPR4 from Arabidopsis thaliana. To facilitate crystallization, T4 lysozyme (T4L) was added to the N-terminus of the AtNPR4 protein. The recombinant T4L-AtNPR4 protein was expressed, purified and crystallized using the sitting-drop and hanging-drop vapour-diffusion methods. The T4L-AtNPR4 crystals have symmetry consistent with space group C2, with unit-cell parameters a = 93.7, b = 85.8, c = 88.2   Å , β = 90 ° and one molecule per asymmetric unit. The best crystal diffracted to a resolution of 2.75   Å . Structure d...
Source: Acta Crystallographica Section F - December 22, 2018 Category: Biochemistry Authors: Yang, Q. Zhang, M. Zheng, J. Jia, Z. Tags: salicylic acid systemic acquired resistance protein purification regulatory mechanism crystal diffraction Arabidopsis thaliana research communications Source Type: research

The redefined DNA-binding domain of human xeroderma pigmentosum complementation group A: production, crystallization and structure solution
Human xeroderma pigmentosum complementation group A (XPA) is a scaffold protein that plays significant roles in DNA-damage verification and in recruiting downstream endonucleases to facilitate the repair of DNA lesions in nucleotide-excision repair. XPA98 – 219 (residues 98 – 219) has been identified as a DNA-binding domain and has been extensively studied in the last two decades. However, the most recent studies have redefined the DNA-binding domain as XPA98 – 239 (residues 98 – 239); it exerts a remarkably higher DNA-binding affinity than XPA98 – 219 and has a binding affinity that is quite ...
Source: Acta Crystallographica Section F - December 22, 2018 Category: Biochemistry Authors: Lian, F.-M. Yang, X. Yang, W. Jiang, Y.-L. Qian, C. Tags: XPA DNA-binding domain nucleotide-excision repair research communications Source Type: research

The structure and activity of the glutathione reductase from Streptococcus pneumoniae
The glutathione reductase (GR) from Streptococcus pneumoniae is a flavoenzyme that catalyzes the reduction of oxidized glutathione (GSSG) to its reduced form (GSH) in the cytoplasm of this bacterium. The maintenance of an intracellular pool of GSH is critical for the detoxification of reactive oxygen and nitrogen species and for intracellular metal tolerance to ions such as zinc. Here, S. pneumoniae GR (SpGR) was overexpressed and purified and its crystal structure determined at 2.56   Å resolution. SpGR shows overall structural similarity to other characterized GRs, with a dimeric structure that includes an antipa...
Source: Acta Crystallographica Section F - December 22, 2018 Category: Biochemistry Authors: Sikanyika, M. Arag ã o, D. McDevitt, C.A. Maher, M.J. Tags: glutathione reductase X-ray crystallography Streptococcus pneumoniae research communications Source Type: research

The X-ray crystal structure of human endothelin 1, a polypeptide hormone regulator of blood pressure
Human endothelin is a 21-amino-acid polypeptide, constrained by two intra-chain disulfide bridges, that is made by endothelial cells. It is the most potent vasoconstrictor in the body and is crucially important in the regulation of blood pressure. It plays a major role in a host of medical conditions, including hypertension, diabetes, stroke and cancer. Endothelin was crystallized 28 years ago in the putative space group P6122, but the structure was never successfully solved by X-ray diffraction. Using X-ray diffraction data from 1992, the structure has now been solved. Assuming a unit cell belonging to space group P61 and...
Source: Acta Crystallographica Section F - December 22, 2018 Category: Biochemistry Authors: McPherson, A. Larson, S.B. Tags: endothelin blood pressure vasoconstrictors polypeptide hormones twinned crystals crystallographic archeology research communications Source Type: research

Never too late for endothelin
(Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - December 22, 2018 Category: Biochemistry Authors: Stanfield, R.L. Tags: endothelin archiving raw data scientific commentaries Source Type: research

Single-particle reconstruction statistics: a diagnostic tool in solving biomolecular structures by cryo-EM
This study revisits the theory of 3D reconstruction and demonstrates how the associated statistics can provide a diagnostic tool to improve SPA. Small numbers of images already give sufficient information on micrograph quality and the amount of data required to reach high resolution. Such feedback allows the microscopist to improve sample-preparation and imaging parameters before committing to extensive data collection. Once a larger data set is available, a B factor can be determined describing the suppression of the signal owing to one or more causes, such as specimen movement, radiation damage, alignment inaccuracy and ...
Source: Acta Crystallographica Section F - December 22, 2018 Category: Biochemistry Authors: Heymann, J.B. Tags: cryo-EM cryo-electron microscopy spectral signal-to-noise ratio single-particle analysis Fourier shell correlation image processing research communications Source Type: research

Survey of the analysis of continuous conformational variability of biological macromolecules by electron microscopy
Single-particle analysis by electron microscopy is a well established technique for analyzing the three-dimensional structures of biological macromolecules. Besides its ability to produce high-resolution structures, it also provides insights into the dynamic behavior of the structures by elucidating their conformational variability. Here, the different image-processing methods currently available to study continuous conformational changes are reviewed. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - December 22, 2018 Category: Biochemistry Authors: Sorzano, C.O.S. Jim é nez, A. Mota, J. Vilas, J.L. Maluenda, D. Mart í nez, M. Ram í rez-Aportela, E. Majtner, T. Segura, J. S á nchez-Garc í a, R. Rancel, Y. del Ca ñ o, L. Conesa, P. Melero, R. Jonic, S. Vargas, J. Cazals, F. Freyberg, Z. Krieger, Tags: electron microscopy image processing continuous heterogeneity single-particle analysis normal-mode analysis research communications Source Type: research

On cross-correlations, averages and noise in electron microscopy
Biological samples are radiation-sensitive and require imaging under low-dose conditions to minimize damage. As a result, images contain a high level of noise and exhibit signal-to-noise ratios that are typically significantly smaller than 1. Averaging techniques, either implicit or explicit, are used to overcome the limitations imposed by the high level of noise. Averaging of 2D images showing the same molecule in the same orientation results in highly significant projections. A high-resolution structure can be obtained by combining the information from many single-particle images to determine a 3D structure. Similarly, a...
Source: Acta Crystallographica Section F - December 21, 2018 Category: Biochemistry Authors: Radermacher, M. Ruiz, T. Tags: image processing signal-to-noise ratio cross-correlation multireference alignment 3D reference-based projection alignment research communications Source Type: research