Crystal structure of the extracellular domain of the receptor-like kinase TMK3 from Arabidopsis thaliana
Transmembrane kinases (TMKs) are members of the plant receptor-like kinase (RLK) family. TMKs are characterized by an extracellular leucine-rich-repeat (LRR) domain, a single transmembrane region and a cytoplasmic kinase domain. TMKs have been shown to act as critical modulators of cell expansion and cell proliferation. Here, the crystal structure of the extracellular domain of TMK3 (TMK3-ECD) was determined to a resolution of 2.06   Å , with an Rwork of 17.69% and an Rfree of 20.58%. Similar to the extracellular domain of TMK1, the TMK3-ECD structure contains two solenoids with 13 LRRs and a non-LRR region (316 &n...
Source: Acta Crystallographica Section F - July 29, 2020 Category: Biochemistry Authors: Chen, H. Kong, Y. Chen, J. Li, L. Li, X. Yu, F. Ming, Z. Tags: transmembrane kinases extracellular domain LRR-RLKs non-LRR region TMK3 protein crystallography research communications Source Type: research

Structural characterization of three noncanonical NTF2-like superfamily proteins: implications for polyketide biosynthesis
Proteins belonging to the NTF2-like superfamily are present in the biosynthetic pathways of numerous polyketide natural products, such as anthracyclins and benzoisochromanequinones. Some have been found to be bona fide polyketide cyclases, but many of them have roles that are currently unknown. Here, the X-ray crystal structures of three NTF2-like proteins of unknown function are reported: those of ActVI-ORFA from Streptomyces coelicolor A3(2) and its homologs Caci_6494, a protein from an uncharacterized biosynthetic cluster in Catenulispora acidiphila, and Aln2 from Streptomyces sp. CM020, a protein in the biosynthetic pa...
Source: Acta Crystallographica Section F - July 29, 2020 Category: Biochemistry Authors: Vuksanovic, N. Zhu, X. Serrano, D.A. Siitonen, V. Mets ä -Ketel ä , M. Melan ç on, C.E. Silvaggi, N.R. Tags: NTF2-like superfamily polyketide cyclases actinorhodin alnumycin secondary metabolism research communications Source Type: research

Structural analysis of a novel substrate-free form of the aminoglycoside 6 ′ -N-acetyltransferase from Enterococcus faecium
In this study, the crystal structure of Ef-AAC(6 ′ )-Ii was determined in a novel substrate-free form. Based on structural analysis, it is proposed that Ef-AAC(6 ′ )-Ii sequentially undergoes conformational selection and induced fit for substrate binding. These results therefore provide a novel viewpoint on the mechanism of action of Ef-AAC(6 ′ )-Ii. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - July 28, 2020 Category: Biochemistry Authors: Jang, H. Kwon, S. Jeong, C.-S. Lee, C.W. Hwang, J. Jung, K.H. Lee, J.H. Park, H.H. Tags: aminoglycoside acetyltransferases Enterococcus faecium acetyl-CoA conformational selection induced fit research communications Source Type: research

Crystal structure of the Rab-binding domain of Rab11 family-interacting protein 2
The small GTPases Rab11, Rab14 and Rab25 regulate membrane trafficking through the recruitment of Rab11 family-interacting proteins (FIPs) to endocytic compartments. FIPs are multi-domain effector proteins that have a highly conserved Rab-binding domain (RBD) at their C-termini. Several structures of complexes of Rab11 with RBDs have previously been determined, including those of Rab11 – FIP2 and Rab11 – FIP3. In addition, the structures of the Rab14 – FIP1 and Rab25 – FIP2 complexes have been determined. All of the RBD structures contain a central parallel coiled coil in the RBD that binds to the s...
Source: Acta Crystallographica Section F - July 28, 2020 Category: Biochemistry Authors: Kearney, A.M. Khan, A.R. Tags: Rab11 family-interacting protein 2 ab initio phasing Rab-binding domain GTPases research communications Source Type: research

Structure of cyclin-dependent kinase 2 (CDK2) in complex with the specific and potent inhibitor CVT-313
CVT-313 is a potent CDK2 inhibitor that was identified by screening a purine-analogue library and is currently in preclinical studies. Since this molecule has the potential to be developed as a CDK2 inhibitor for cancer therapy, the potency of CVT-313 to bind and stabilize CDK2 was evaluated, together with its ability to inhibit aberrant cell proliferation. CVT-313 increased the melting temperature of CDK2 by 7 ° C in thermal stabilization studies, thus indicating its protein-stabilizing effect. CVT-313 inhibited the growth of human lung carcinoma cell line A549 in a dose-dependent manner, with an IC50 of 1.2   &micr...
Source: Acta Crystallographica Section F - July 28, 2020 Category: Biochemistry Authors: Talapati, S.R. Nataraj, V. Pothuganti, M. Gore, S. Ramachandra, M. Antony, T. More, S.S. Krishnamurthy, N.R. Tags: CDK2 cyclin-dependent kinase 2 CVT-313 crystal structure X-ray crystallography cancer therapy research communications Source Type: research

Crystal structure of GH30-7 endoxylanase C from the filamentous fungus Talaromyces cellulolyticus
In this study, the crystal structure of a TcXyn30C mutant which lacks the CBM1 domain was determined at 1.65   Å resolution. The structure of the active site of TcXyn30C was compared with that of the bifunctional GH30-7 xylanase B from T. cellulolyticus (TcXyn30B), which exhibits glucuronoxylanase and xylobiohydrolase activities. The results revealed that TcXyn30C has a conserved structural feature for recognizing the 4-O-methyl- α -d-glucuronic acid (MeGlcA) substituent in subsite − 2b. Additionally, the results demonstrated that Phe47 contributes significantly to catalysis by TcXyn30C. Phe47 is locate...
Source: Acta Crystallographica Section F - July 28, 2020 Category: Biochemistry Authors: Nakamichi, Y. Fujii, T. Watanabe, M. Matsushika, A. Inoue, H. Tags: crystal structure endoxylanases glucuronoxylanases glycoside hydrolase family 30 Talaromyces cellulolyticus research communications Source Type: research

Expression, purification and crystal structure determination of a ferredoxin reductase from the actinobacterium Thermobifida fusca
The ferredoxin reductase FdR9 from Thermobifida fusca, a member of the oxygenase-coupled NADH-dependent ferredoxin reductase (FNR) family, catalyses electron transfer from NADH to its physiological electron acceptor ferredoxin. It forms part of a putative three-component cytochrome P450 monooxygenase system in T. fusca comprising CYP222A1 and the [3Fe – 4S]-cluster ferredoxin Fdx8 as well as FdR9. Here, FdR9 was overexpressed and purified and its crystal structure was determined at 1.9   Å resolution. The overall structure of FdR9 is similar to those of other members of the FNR family and is composed of an FA...
Source: Acta Crystallographica Section F - July 28, 2020 Category: Biochemistry Authors: Rodriguez Buitrago, J.A. Kl ü nemann, T. Blankenfeldt, W. Schallmey, A. Tags: ferredoxin reductase Thermobifida fusca cytochrome P450 research communications Source Type: research

Crystal structure of the FYCO1 RUN domain suggests possible interfaces with small GTPases
FYCO1 is a multidomain adaptor protein that plays an important role in autophagy by mediating the kinesin-dependent microtubule plus-end-directed transport of autophagosomes. FYCO1 contains a RUN domain, which is hypothesized to function as a specific effector for members of the Ras superfamily of small GTPases, but its role has not been well characterized and its interaction partner(s) have not been identified. Here, the crystal structure of the FYCO1 RUN domain was determined at 1.3   Å resolution. The overall structure of the FYCO1 RUN domain was similar to those of previously reported RUN domains. Detailed stru...
Source: Acta Crystallographica Section F - July 28, 2020 Category: Biochemistry Authors: Sakurai, S. Shimizu, T. Ohto, U. Tags: X-ray crystallography autophagy FYCO1 RUN domain small GTPase binding research communications Source Type: research

Innovation versus practice in biological crystallization
(Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - July 1, 2020 Category: Biochemistry Authors: Newman, J. van Raaij, M.J. Tags: crystallization innovation editorial Source Type: research

Proline/alanine-rich sequence (PAS) polypeptides as an alternative to PEG precipitants for protein crystallization
Proline/alanine-rich sequence (PAS) polypeptides represent a novel class of biosynthetic polymers comprising repetitive sequences of the small proteinogenic amino acids l-proline, l-alanine and/or l-serine. PAS polymers are strongly hydrophilic and highly soluble in water, where they exhibit a natively disordered conformation without any detectable secondary or tertiary structure, similar to polyethylene glycol (PEG), which constitutes the most widely applied precipitant for protein crystallization to date. To investigate the potential of PAS polymers for structural studies by X-ray crystallography, two proteins that were ...
Source: Acta Crystallographica Section F - July 1, 2020 Category: Biochemistry Authors: Schiefner, A. Walser, R. Gebauer, M. Skerra, A. Tags: disordered polypeptide PASylation protein precipitant polyamino acid polyethylene glycol proline/alanine-rich sequence protein crystallization methods communications Source Type: research

A proposed carbon-utilization and virulence protein A, CuvA (Rv1422), from Mycobacterium tuberculosis H37Rv: crystallization, X-ray diffraction analysis and ligand binding
Mycobacterium tuberculosis possesses the ability to undergo physiological adaptations in order to persist during the prolonged course of infection despite the active immune response of the host and in order to overcome multiple environmental changes. Previous studies have proposed that M. tuberculosis CuvA (Rv1422; MtCuvA) might play a critical role in the adaptation of the bacterium to environmental changes, such as nutrient utilization and alteration of the growth rate. However, the detailed function of MtCuvA still remains unclear owing to a lack of structural information. To better understand its role in host adaptatio...
Source: Acta Crystallographica Section F - July 1, 2020 Category: Biochemistry Authors: Jeong, Y.C. Lee, K.S. Tags: Mycobacterium tuberculosis CuvA (Rv1422) bacterial adaptation nutrient utilization cell-wall precursor components research communications Source Type: research

Structural characterization of borneol dehydrogenase from Pseudomonas sp. TCU-HL1
During the microbial degradation of borneol, a bicyclic plant monoterpene, it is first converted into camphor by borneol dehydrogenase (BDH) and then enters a known camphor-degradation pathway. Previously, a recombinant Pseudomonas BDH was found in inclusion bodies when expressed in Escherichia coli. After refolding, it was still unstable and was difficult to concentrate. Here, the protein-expression conditions were improved by changing the medium from lysogeny broth to Terrific Broth, yielding a soluble form of the enzyme with higher activity. The protein was crystallized and its 3D structure was determined by X-ray diffr...
Source: Acta Crystallographica Section F - July 1, 2020 Category: Biochemistry Authors: Khine, A.A. Chen, H.-P. Huang, K.-F. Ko, T.-P. Tags: plant terpenoids camphor oxidoreductases Rossmann fold NAD expression medium protein solubility research communications Source Type: research

Tartryl-CoA inhibits succinyl-CoA synthetase
Succinyl-CoA synthetase (SCS) catalyzes the only substrate-level phosphorylation step in the tricarboxylic acid cycle. Human GTP-specific SCS (GTPSCS), an α β -heterodimer, was produced in Escherichia coli. The purified protein crystallized from a solution containing tartrate, CoA and magnesium chloride, and a crystal diffracted to 1.52   Å resolution. Tartryl-CoA was discovered to be bound to GTPSCS. The CoA portion lies in the amino-terminal domain of the α -subunit and the tartryl end extends towards the catalytic histidine residue. The terminal carboxylate binds to the phosphate-binding site of...
Source: Acta Crystallographica Section F - July 1, 2020 Category: Biochemistry Authors: Huang, J. Fraser, M.E. Tags: thioesters catalysis tricarboxylic acid cycle succinyl-CoA synthetase research communications Source Type: research

Crystal structure of the catalytic subunit of bovine pyruvate dehydrogenase phosphatase
Mammalian pyruvate dehydrogenase (PDH) activity is tightly regulated by phosphorylation and dephosphorylation, which is catalyzed by PDH kinase isomers and PDH phosphatase isomers, respectively. PDH phosphatase isomer 1 (PDP1) is a heterodimer consisting of a catalytic subunit (PDP1c) and a regulatory subunit (PDP1r). Here, the crystal structure of bovine PDP1c determined at 2.1   Å resolution is reported. The crystals belonged to space group P3221, with unit-cell parameters a = b = 75.3, c = 173.2   Å . The structure was solved by molecular-replacement methods and refined to a final R factor of 21.9% (Rfre...
Source: Acta Crystallographica Section F - July 1, 2020 Category: Biochemistry Authors: Guo, Y. Qiu, W. Roche, T.E. Hackert, M.L. Tags: pyruvate dehydrogenase phosphatase cis-peptide research communications Source Type: research

Novel carbohydrate-recognition mode of the invertebrate C-type lectin SPL-1 from Saxidomus purpuratus revealed by the GlcNAc-complex crystal in the presence of Ca2+
The C-type lectins SPL-1 and SPL-2 from the bivalve Saxidomus purpuratus are composed of A and B chains and of two B chains, respectively. They bind specific carbohydrates containing acetamido groups, such as N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc), in a Ca2+-independent manner. Unlike ordinary C-type lectins, which require Ca2+ ions for carbohydrate recognition, these lectins recognize specific carbohydrates mainly through interactions with the acetamido group without Ca2+ ions, even though Ca2+ enhances the binding affinity of these lectins, especially SPL-1. In the present study, the crystal stru...
Source: Acta Crystallographica Section F - June 5, 2020 Category: Biochemistry Authors: Unno, H. Higuchi, S. Goda, S. Hatakeyama, T. Tags: lectins bivalve C-type lectin Saxidomus purpuratus research communications Source Type: research

Crystal structure of the SH3 domain of growth factor receptor-bound protein 2
This study presents the crystal structure of the N-terminal SH3 (SH3N) domain of growth factor receptor-bound protein 2 (Grb2) at 2.5   Å resolution. Grb2 is a small (215-amino-acid) adaptor protein that is widely expressed and involved in signal transduction/cell communication. The crystal structure of full-length Grb2 has previously been reported (PDB entry 1gri). The structure of the isolated SH3N domain is consistent with the full-length structure. The structure of the isolated SH3N domain was solved at a higher resolution (2.5   Å compared with 3.1   Å for the previously deposited structure) an...
Source: Acta Crystallographica Section F - June 5, 2020 Category: Biochemistry Authors: Bolgov, A. Korban, S. Luzik, D. Zhemkov, V. Kim, M. Rogacheva, O. Bezprozvanny, I. Tags: SH3 domain Grb2 adaptor protein growth factor receptor-bound protein 2 research communications Source Type: research

Enhanced X-ray diffraction of in vivo-grown μ NS crystals by viscous jets at XFELs
μ NS is a 70   kDa major nonstructural protein of avian reoviruses, which cause significant economic losses in the poultry industry. They replicate inside viral factories in host cells, and the μ NS protein has been suggested to be the minimal viral factor required for factory formation. Thus, determining the structure of μ NS is of great importance for understanding its role in viral infection. In the study presented here, a fragment consisting of residues 448 – 605 of μ NS was expressed as an EGFP fusion protein in Sf9 insect cells. EGFP- μ NS(448 – 605) crystallization in Sf9 cells was monit...
Source: Acta Crystallographica Section F - May 29, 2020 Category: Biochemistry Authors: Nagaratnam, N. Tang, Y. Botha, S. Saul, J. Li, C. Hu, H. Zaare, S. Hunter, M. Lowry, D. Weierstall, U. Zatsepin, N. Spence, J.C.H. Qiu, J. LaBaer, J. Fromme, P. Martin-Garcia, J.M. Tags: μ NS avian reovirus in vivo crystallization high-viscosity jets serial crystallography X-ray free-electron lasers methods communications Source Type: research

Expression, purification, crystallization and X-ray diffraction studies of a novel root-induced secreted protein from Trichoderma virens
Small secreted cysteine-rich proteins (SSCPs) from fungi play an important role in fungi – host interactions. The plant-beneficial fungi Trichoderma spp. are in use worldwide as biocontrol agents and protect the host plant from soil-borne as well as foliar pathogens. Recently, a novel SSCP, Tsp1, has been identified in the secreted protein pool of T. virens and is overinduced upon its interaction with the roots of the maize plant. The protein was observed to be well conserved in the Ascomycota division of fungi, and its homologs are present in many plant-pathogenic fungi such as Fusarium oxysporum and Magnaporthe ory...
Source: Acta Crystallographica Section F - May 29, 2020 Category: Biochemistry Authors: Bansal, R. Mistry, H.U. Mukherjee, P.K. Gupta, G.D. Tags: Trichoderma virens effector secreted cysteine-rich proteins protein crystallization research communications Source Type: research

Structure of heme d1-free cd1 nitrite reductase NirS
A key step in anaerobic nitrate respiration is the reduction of nitrite to nitric oxide, which is catalysed by the cd1 nitrite reductase NirS in, for example, the Gram-negative opportunistic pathogen Pseudomonas aeruginosa. Each subunit of this homodimeric enzyme consists of a cytochrome c domain and an eight-bladed β -propeller that binds the uncommon isobacteriochlorin heme d1 as an essential part of its active site. Although NirS has been well studied mechanistically and structurally, the focus of previous studies has been on the active heme d1-bound form. The heme d1-free form of NirS reported here, which represen...
Source: Acta Crystallographica Section F - May 29, 2020 Category: Biochemistry Authors: Kl ü nemann, T. Blankenfeldt, W. Tags: cd1 nitrite reductase NirS heme d1 research communications Source Type: research

Crystal structures of the GH18 domain of the bifunctional peroxiredoxin – chitinase CotE from Clostridium difficile
CotE is a coat protein that is present in the spores of Clostridium difficile, an obligate anaerobic bacterium and a pathogen that is a leading cause of antibiotic-associated diarrhoea in hospital patients. Spores serve as the agents of disease transmission, and CotE has been implicated in their attachment to the gut epithelium and subsequent colonization of the host. CotE consists of an N-terminal peroxiredoxin domain and a C-terminal chitinase domain. Here, a C-terminal fragment of CotE comprising residues 349 – 712 has been crystallized and its structure has been determined to reveal a core eight-stranded β -...
Source: Acta Crystallographica Section F - May 29, 2020 Category: Biochemistry Authors: Whittingham, J.L. Hanai, S. Brannigan, J.A. Ferreira, W.T. Dodson, E.J. Turkenburg, J.P. Cartwright, J. Cutting, S.M. Wilkinson, A.J. Tags: Clostridium difficile spores CotE glycosyl hydrolase 3D domain swapping research communications Source Type: research

Crystallization and X-ray analysis of Borrelia burgdorferi β -barrel assembly machinery A
Mitochondria, chloroplasts and several species of bacteria have outer membrane proteins (OMPs) that perform many essential biological functions. The β -barrel assembly machinery (BAM) complex is one of the OMPs of Borrelia burgdorferi, the pathogenic spirochete that causes Lyme disease, and its BamA component (BbBamA) includes a C-terminal β -barrel domain and five N-terminal periplasmic polypeptide-transport-associated (POTRA) domains, which together perform a central transport function. In the current work, the production, crystallization and X-ray analysis of the three N-terminal POTRA domains of BbBamA (BbBam...
Source: Acta Crystallographica Section F - May 29, 2020 Category: Biochemistry Authors: Dong, S. Chu, H. Wen, K. Yu, Q. Li, H. Wang, C. Qin, X. Tags: Borrelia burgdorferi BbBamA-POTRA P1 – P3 crystallization research communications Source Type: research

Crystal structure of gluconate 5-dehydrogenase from Lentibacter algarum
Gluconate 5-dehydrogenase (Ga5DH; EC 1.1.1.69) from Lentibacter algarum (LaGa5DH) was recombinantly expressed in Escherichia coli and purified to homogeneity. The protein was crystallized and the crystal structure was solved at 2.1   Å resolution. The crystal belonged to the monoclinic system, with space group P1 and unit-cell parameters a = 55.42, b   =   55.48, c   =   79.16   Å , α = 100.51, β   =   105.66, γ = 97.99 ° . The structure revealed LaGaDH to be a tetramer, with each subunit consisting of six α -helices and three antiparallel β -hai...
Source: Acta Crystallographica Section F - April 29, 2020 Category: Biochemistry Authors: Tian, D. Fu, X. Cao, W. Yuan, H. Tags: crystal structure gluconate 5-dehydrogenase Lentibacter algarum research communications Source Type: research

Crystallographic snapshots of the EF-hand protein MCFD2 complexed with the intracellular lectin ERGIC-53 involved in glycoprotein transport
The transmembrane intracellular lectin ER – Golgi intermediate compartment protein 53 (ERGIC-53) and the soluble EF-hand multiple coagulation factor deficiency protein 2 (MCFD2) form a complex that functions as a cargo receptor, trafficking various glycoproteins between the endoplasmic reticulum (ER) and the Golgi apparatus. It has been demonstrated that the carbohydrate-recognition domain (CRD) of ERGIC-53 (ERGIC-53CRD) interacts with N-linked glycans on cargo glycoproteins, whereas MCFD2 recognizes polypeptide segments of cargo glycoproteins. Crystal structures of ERGIC-53CRD complexed with MCFD2 and mannosyl oligo...
Source: Acta Crystallographica Section F - April 29, 2020 Category: Biochemistry Authors: Satoh, T. Nishio, M. Suzuki, K. Yagi-Utsumi, M. Kamiya, Y. Mizushima, T. Kato, K. Tags: cargo receptor conformational polymorphism crystal packing glycoprotein transport intracellular lectin MCFD2 ERGIC-53 research communications Source Type: research

Crystal structure of the nucleoid-associated protein Fis (PA4853) from Pseudomonas aeruginosa
Factor for inversion stimulation (Fis) is a versatile bacterial nucleoid-associated protein that can directly bind and bend DNA to influence DNA topology. It also plays crucial roles in regulating bacterial virulence factors and in optimizing bacterial adaptation to various environments. Fis from Pseudomonas aeruginosa (PA4853, referred to as PaFis) has recently been found to be required for virulence by regulating the expression of type III secretion system (T3SS) genes. PaFis can specifically bind to the promoter region of exsA, which functions as a T3SS master regulator, to regulate its expression and plays an essential...
Source: Acta Crystallographica Section F - April 29, 2020 Category: Biochemistry Authors: Zhou, J. Gao, Z. Zhang, H. Dong, Y. Tags: nucleoid-associated protein Fis DNA-binding protein crystal structure helix – turn helix motif Pseudomonas aeruginosa factor for inversion stimulation research communications Source Type: research

Structure of the 4-hydroxy-tetrahydrodipicolinate synthase from the thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV and the phylogeny of the aminotransferase pathway
The enzyme 4-hydroxy-tetrahydrodipicolinate synthase (DapA) is involved in the production of lysine and precursor molecules for peptidoglycan synthesis. In a multistep reaction, DapA converts pyruvate and l-aspartate-4-semialdehyde to 4-hydroxy-2,3,4,5-tetrahydrodipicolinic acid. In many organisms, lysine binds allosterically to DapA, causing negative feedback, thus making the enzyme an important regulatory component of the pathway. Here, the 2.1   Å resolution crystal structure of DapA from the thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV is reported. The enzyme crystallized as a contaminant ...
Source: Acta Crystallographica Section F - April 28, 2020 Category: Biochemistry Authors: Schmitz, R.A. Dietl, A. M ü ller, M. Berben, T. Op den Camp, H.J.M. Barends, T.R.M. Tags: 4-hydroxy-tetrahydrodipicolinate synthase Methylacidiphilum fumariolicum SolV methanotroph aminotransferase pathway research communications Source Type: research

Characterization of the Pseudomonas aeruginosa T6SS PldB immunity proteins PA5086, PA5087 and PA5088 explains a novel stockpiling mechanism
The bacterial type VI secretion system (T6SS) secretes many toxic effectors to gain advantage in interbacterial competition and for eukaryotic host infection. The cognate immunity proteins of these effectors protect bacteria from their own effectors. PldB is a T6SS trans-kingdom effector in Pseudomonas aeruginosa that can infect both prokaryotic and eukaryotic cells. Three proteins, PA5086, PA5087 and PA5088, are employed to suppress the toxicity of PldB-family proteins. The structures of PA5087 and PA5088 have previously been reported, but the identification of further distinctions between these immunity proteins is neede...
Source: Acta Crystallographica Section F - April 28, 2020 Category: Biochemistry Authors: Wen, H. Geng, Z. Gao, Z. She, Z. Dong, Y. Tags: X-ray crystallography T6SS immunity proteins stockpiling mechanism electrostatic potential surface PA5086 Pseudomonas aeruginosa type VI secretion system research communications Source Type: research

Structure of a single-chain H2A/H2B dimer
Chromatin is the complex assembly of nucleic acids and proteins that makes up the physiological form of the eukaryotic genome. The nucleosome is the fundamental repeating unit of chromatin, and is composed of ∼ 147   bp of DNA wrapped around a histone octamer formed by two copies of each core histone: H2A, H2B, H3 and H4. Prior to nucleosome assembly, and during histone eviction, histones are typically assembled into soluble H2A/H2B dimers and H3/H4 dimers and tetramers. A multitude of factors interact with soluble histone dimers and tetramers, including chaperones, importins, histone-modifying enzymes and chromatin-...
Source: Acta Crystallographica Section F - April 28, 2020 Category: Biochemistry Authors: Warren, C. Bonanno, J.B. Almo, S.C. Shechter, D. Tags: histones chromatin H2A/H2B dimer Xenopus laevis crystallography research communications Source Type: research

Crystal structure of Arabidopsis thaliana casein kinase 2 α 1
Casein kinase 2 (CK2) is a ubiquitous pleiotropic enzyme that is highly conserved across eukaryotic kingdoms. CK2 is singular amongst kinases as it is highly rigid and constitutively active. Arabidopsis thaliana is widely used as a model system in molecular plant research; the biological functions of A.   thaliana CK2 are well studied in vivo and many of its substrates have been identified. Here, crystal structures of the α subunit of A. thaliana CK2 in three crystal forms and of its complex with the nonhydrolyzable ATP analog AMppNHp are presented. While the C-lobe of the enzyme is highly rigid, structural plas...
Source: Acta Crystallographica Section F - April 6, 2020 Category: Biochemistry Authors: Demulder, M. De Veylder, L. Loris, R. Tags: casein kinase CK2 structural flexibility ATP binding Arabidopsis thaliana research communications Source Type: research

Structure of the Mycobacterium smegmatis α -maltose-1-phosphate synthase GlgM
Mycobacterium tuberculosis produces glycogen (also known as α -glucan) to help evade human immunity. This pathogen uses the GlgE pathway to generate glycogen rather than the more well known glycogen synthase GlgA pathway, which is absent in this bacterium. Thus, the building block for this glucose polymer is α -maltose-1-phosphate rather than an NDP-glucose donor. One of the routes to α -maltose-1-phosphate is now known to involve the GlgA homologue GlgM, which uses ADP-glucose as a donor and α -glucose-1-phosphate as an acceptor. To help compare GlgA (a GT5 family member) with GlgM enzymes (GT4 fam...
Source: Acta Crystallographica Section F - April 3, 2020 Category: Biochemistry Authors: Syson, K. Stevenson, C.E.M. Lawson, D.M. Bornemann, S. Tags: GlgM α -maltose-1-phosphate synthase glycosyltransferase glycogen -glucan research communications Source Type: research

Rv0100, a proposed acyl carrier protein in Mycobacterium tuberculosis: expression, purification and crystallization. Corrigendum
The true identity of the protein found in the crystals reported by Bondoc et al. [(2019), Acta Cryst. F75, 646 – 651] is given. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 2, 2020 Category: Biochemistry Authors: Bondoc, J.M.G. Gutka, H.J. Almutairi, M.M. Patwell, R. Rutter, M.W. Wolf, N.M. Samudrala, R. Mehboob, S. Dementiev, A. Abad-Zapatero, C. Movahedzadeh, F. Tags: acyl carrier protein Mycobacterium tuberculosis Mycobacterium smegmatis Rv0100 corrigendum addenda and errata Source Type: research

Visualizing an unseen enemy; mobilizing structural biology to counter COVID-19
(Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 1, 2020 Category: Biochemistry Authors: Baker, E.N. Tags: COVID-19 coronaviruses SARS-CoV-2 editorial Source Type: research

Crystal structure of monomeric Amuc_1100 from Akkermansia muciniphila
This study provides a structural basis for the elucidation of the molecular mechanism of Amuc_1100. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 1, 2020 Category: Biochemistry Authors: Mou, L. Peng, X. Chen, Y. Xiao, Q. Liao, H. Liu, M. Guo, L. Liu, Y. Zhang, X. Deng, D. Tags: Akkermansia muciniphila type IV pili Amuc_1100 gut microbes crystal structure research communications Source Type: research

Structural characterization of human O-phosphoethanolamine phospho-lyase
Human O-phosphoethanolamine phospho-lyase (hEtnppl; EC 4.2.3.2) is a pyridoxal 5 ′ -phosphate-dependent enzyme that catalyzes the degradation of O-phosphoethanolamine (PEA) into acetaldehyde, phosphate and ammonia. Physiologically, the enzyme is involved in phospholipid metabolism, as PEA is the precursor of phosphatidylethanolamine in the CDP-ethanolamine (Kennedy) pathway. Here, the crystal structure of hEtnppl in complex with pyridoxamine 5 ′ -phosphate was determined at 2.05   Å resolution by molecular replacement using the structure of A1RDF1 from Arthrobacter aurescens TC1 (PDB entry 5g4i) as the ...
Source: Acta Crystallographica Section F - April 1, 2020 Category: Biochemistry Authors: Vettraino, C. Peracchi, A. Donini, S. Parisini, E. Tags: O-phosphoethanolamine phospho-lyase lyases PMP crystal structure research communications Source Type: research

Crystal structure of Arabidopsis thaliana neutral invertase 2
The metabolism of sucrose is of crucial importance for life on Earth. In plants, enzymes called invertases split sucrose into glucose and fructose, contributing to the regulation of metabolic fluxes. Invertases differ in their localization and pH optimum. Acidic invertases present in plant cell walls and vacuoles belong to glycoside hydrolase family 32 (GH32) and have an all- β structure. In contrast, neutral invertases are located in the cytosol and organelles such as chloroplasts and mitochondria. These poorly understood enzymes are classified into a separate GH100 family. Recent crystal structures of the closely re...
Source: Acta Crystallographica Section F - March 3, 2020 Category: Biochemistry Authors: Tarkowski, Ł .P. Tsirkone, V.G. Osipov, E.M. Beelen, S. Lammens, W. Vergauwen, R. Van den Ende, W. Strelkov, S.V. Tags: GH100 neutral invertase Arabidopsis thaliana crystal structure research communications Source Type: research

Structural insights into the catalytic mechanism of Bacillus subtilis BacF
The nonribosomal biosynthesis of the dipeptide antibiotic bacilysin is achieved by the concerted action of multiple enzymes in the Bacillus subtilis bac operon. BacF (YwfG), encoded by the bacF gene, is a fold type I pyridoxal 5-phosphate (PLP)-dependent stereospecific transaminase. Activity assays with l-phenylalanine and 4-hydroxyphenylpyruvic acid (4HPP), a chemical analogue of tetrahydrohydroxyphenylpyruvic acid (H4HPP), revealed stereospecific substrate preferences, a finding that was consistent with previous reports on the role of this enzyme in bacilysin synthesis. The crystal structure of this dimeric enzyme was de...
Source: Acta Crystallographica Section F - March 3, 2020 Category: Biochemistry Authors: Deshmukh, A. Gopal, B. Tags: transaminases external aldimine stereospecificity enzyme catalysis bacilysin synthesis research communications Source Type: research

The phospholipase A effector PlaA from Legionella pneumophila: expression, purification and crystallization
Legionella pneumophila encodes an extracellular secreted phospholipase A named PlaA that is translocated by the type II secretion system. It plays an essential role in maintaining the integrity of Legionella-containing vacuoles in L. pneumophila pathogenesis. Here, it is shown that PlaA has a main lysophospholipase activity to hydrolyze fatty-acyl groups in lysophospholipids. Although it has a very low phospholipase A activity to catalyze the hydrolysis of fatty-acyl groups in phospholipids, PlaA can bind phospholipids such as 1,2-dipalmitoylphosphatidylcholine with a dissociation constant of 11.1   µ M. Sequence-a...
Source: Acta Crystallographica Section F - March 2, 2020 Category: Biochemistry Authors: Qu, X. Song, X. Zhang, N. Ma, J. Ge, H. Tags: phospholipase A Legionella pneumophila effector proteins research communications Source Type: research

Structure determination of the human TRPV1 ankyrin-repeat domain under nonreducing conditions
TRPV1, a member of the transient receptor potential (TRP) channels family, has been found to be involved in redox sensing. The crystal structure of the human TRPV1 ankyrin-repeat domain (TRPV1-ARD) was determined at 4.5   Å resolution under nonreducing conditions. This is the first report of the crystal structure of a ligand-free form of TRPV1-ARD and in particular of the human homologue. The structure showed a unique conformation in finger loop 3 near Cys258, which is most likely to be involved in inter-subunit disulfide-bond formation. Also, in human TRPV1-ARD it was possible for solvent to access Cys258. This st...
Source: Acta Crystallographica Section F - March 2, 2020 Category: Biochemistry Authors: Tanaka, M. Hayakawa, K. Ogawa, N. Kurokawa, T. Kitanishi, K. Ite, K. Matsui, T. Mori, Y. Unno, M. Tags: TRPV1 akyrin-repeat domain human nonreducing conditions research communications Source Type: research

Engineering the Fab fragment of the anti-IgE omalizumab to prevent Fab crystallization and permit IgE-Fc complex crystallization
Immunoglobulin E (IgE) plays a central role in the allergic response, in which cross-linking of allergen by Fc ∊ RI-bound IgE triggers mast cell and basophil degranulation and the release of inflammatory mediators. The high-affinity interaction between IgE and Fc ∊ RI is a long-standing target for therapeutic intervention in allergic disease. Omalizumab is a clinically approved anti-IgE monoclonal antibody that binds to free IgE, also with high affinity, preventing its interaction with Fc ∊ RI. All attempts to crystallize the pre-formed complex between the omalizumab Fab and the Fc region of IgE (IgE-Fc), to understa...
Source: Acta Crystallographica Section F - March 2, 2020 Category: Biochemistry Authors: Mitropoulou, A.N. Ceska, T. Heads, J.T. Beavil, A.J. Henry, A.J. McDonnell, J.M. Sutton, B.J. Davies, A.M. Tags: omalizumab allergy Fab immunoglobulin E antibody protein engineering X-ray crystallography research communications Source Type: research

Structural and biochemical characterization of mitochondrial citrate synthase 4 from Arabidopsis thaliana
Citrate synthase (CS) catalyzes the conversion of oxaloacetate and acetyl coenzyme A into citrate and coenzyme A in the mitochondrial tricarboxylic acid (TCA) cycle. In plants, mitochondrial metabolism, including the TCA cycle, occurs in interaction with photosynthetic metabolism. The controlled regulation of several enzymes in the TCA cycle, such as CS, is important in plants. Here, the first crystal structure of a plant mitochondrial CS, CSY4 from Arabidopsis thaliana (AtCSY4), has been determined. Structural comparison of AtCSY4 with mitochondrial CSs revealed a high level of similarity. Inhibition analysis showed a sim...
Source: Acta Crystallographica Section F - March 1, 2020 Category: Biochemistry Authors: Nishio, K. Mizushima, T. Tags: citrate synthase mitochondria TCA cycle Arabidopsis thaliana X-ray crystallography research communications Source Type: research

The crystal structure of the catalytic domain of tau tubulin kinase 2 in complex with a small-molecule inhibitor
Tau proteins play an important role in the proper assembly and function of neurons. Hyperphosphorylation of tau by kinases such as tau tubulin kinase (TTBK) has been hypothesized to cause the aggregation of tau and the formation of neurofibrillary tangles (NFTs) that lead to the destabilization of microtubules, thereby contributing to neurodegenerative diseases such as Alzheimer's disease (AD). There are two TTBK isoforms with highly homologous catalytic sites but with distinct tissue distributions, tau phosphorylation patterns and loss-of-function effects. Inhibition of TTBK1 reduces the levels of NFT formation involved i...
Source: Acta Crystallographica Section F - March 1, 2020 Category: Biochemistry Authors: Marcotte, D.J. Spilker, K.A. Wen, D. Hesson, T. Patterson, T.A. Kumar, P.R. Chodaparambil, J.V. Tags: TTBK2 kinase domain tau proteins Alzheimer's disease research communications Source Type: research

High-resolution crystal structures of Escherichia coli FtsZ bound to GDP and GTP
Bacterial cytokinesis is mediated by the Z-ring, which is formed by the prokaryotic tubulin homolog FtsZ. Recent data indicate that the Z-ring is composed of small patches of FtsZ protofilaments that travel around the bacterial cell by treadmilling. Treadmilling involves a switch from a relaxed (R) state, favored for monomers, to a tense (T) conformation, which is favored upon association into filaments. The R conformation has been observed in numerous monomeric FtsZ crystal structures and the T conformation in Staphylococcus aureus FtsZ crystallized as assembled filaments. However, while Escherichia coli has served as a m...
Source: Acta Crystallographica Section F - February 5, 2020 Category: Biochemistry Authors: Schumacher, M.A. Ohashi, T. Corbin, L. Erickson, H.P. Tags: FtsZ cytokinesis treadmilling protofilaments Z-ring research communications Source Type: research

Crystal structures of the cell-division protein FtsZ from Klebsiella pneumoniae and Escherichia coli
In this study, crystal structures of the enzymatic domains of FtsZ from Klebsiella pneumoniae (KpFtsZ) and Escherichia coli (EcFtsZ) were determined at 1.75 and 2.50   Å resolution, respectively. Both FtsZs form straight protofilaments in the crystals, and the two structures adopted relaxed (R) conformations. The T3 loop, which is   involved in GTP/GDP binding and FtsZ assembly/disassembly, adopted a unique open conformation in KpFtsZ, while the T3 loop of EcFtsZ was partially disordered. The crystal structure of EcFtsZ can explain the results from previous functional analyses using EcFtsZ mutants. (Source: Ac...
Source: Acta Crystallographica Section F - February 5, 2020 Category: Biochemistry Authors: Yoshizawa, T. Fujita, J. Terakado, H. Ozawa, M. Kuroda, N. Tanaka, S. Uehara, R. Matsumura, H. Tags: cell division FtsZ Escherichia coli Klebsiella pneumoniae research communications Source Type: research

An unexpected co-crystal structure of the calpain PEF(S) domain with Hfq reveals a potential chaperone function of Hfq
Calpain is a Ca2+-activated, heterodimeric cysteine protease consisting of a large catalytic subunit and a small regulatory subunit. Dysregulation of this enzyme is involved in a range of pathological conditions such as cancer, Alzheimer's disease and rheumatoid arthritis, and thus calpain I is a drug target with potential therapeutic applications. Difficulty in the production of this enzyme has hindered structural and functional investigations in the past, although heterodimeric calpain I can be generated by Escherichia coli expression in low yield. Here, an unexpected structure discovered during crystallization trials of...
Source: Acta Crystallographica Section F - February 5, 2020 Category: Biochemistry Authors: Cresser-Brown, J. Rizkallah, P. Jin, Y. Roth, C. Miller, D.J. Allemann, R.K. Tags: calpain expression Hfq chaperones cysteine protease research communications Source Type: research

The thermodynamic profile and molecular interactions of a C(9)-cytisine derivative-binding acetylcholine-binding protein from Aplysia californica
Cytisine, a natural product with high affinity for clinically relevant nicotinic acetylcholine receptors (nAChRs), is used as a smoking-cessation agent. The compound displays an excellent clinical profile and hence there is an interest in derivatives that may be further improved or find use in the treatment of other conditions. Here, the binding of a cytisine derivative modified by the addition of a 3-(hydroxypropyl) moiety (ligand 4) to Aplysia californica acetylcholine-binding protein (AcAChBP), a surrogate for nAChR orthosteric binding sites, was investigated. Isothermal titration calorimetry revealed that the favorable...
Source: Acta Crystallographica Section F - February 3, 2020 Category: Biochemistry Authors: Davis, S. Rego Campello, H. Gallagher, T. Hunter, W.N. Tags: acetylcholine-binding protein crystal structure cytisine ligand-gated ion channel nicotine research communications Source Type: research

Conformational flexibility within the small domain of human serine racemase
Serine racemase (SR) is a pyridoxal 5 ′ -phosphate (PLP)-containing enzyme that converts l-serine to d-serine, an endogenous co-agonist for the N-methyl-d-aspartate receptor (NMDAR) subtype of glutamate ion channels. SR regulates d-serine levels by the reversible racemization of l-serine to d-serine, as well as the catabolism of serine by α , β -elimination to produce pyruvate. The modulation of SR activity is therefore an attractive therapeutic approach to disorders associated with abnormal glutamatergic signalling since it allows an indirect modulation of NMDAR function. In the present study, a 1.89   ...
Source: Acta Crystallographica Section F - February 3, 2020 Category: Biochemistry Authors: Koulouris, C.R. Bax, B.D. Atack, J.R. Roe, S.M. Tags: serine racemase d-serine NMDA receptors drug design domain structure subdomain ligand-induced reorientation research communications Source Type: research

Structure of the AAVhu.37 capsid by cryoelectron microscopy
Adeno-associated viruses (AAVs) are used as in vivo gene-delivery vectors in gene-therapy products and have been heavily investigated for numerous indications. Over 100 naturally occurring AAV serotypes and variants have been isolated from primate samples. Many reports have described unique properties of these variants (for instance, differences in potency, target cell or evasion of the immune response), despite high amino-acid sequence conservation. AAVhu.37 is of interest for clinical applications owing to its proficient transduction of the liver and central nervous system. The sequence identity of the AAVhu.37 VP1 to th...
Source: Acta Crystallographica Section F - February 3, 2020 Category: Biochemistry Authors: Kaelber, J.T. Yost, S.A. Webber, K.A. Firlar, E. Liu, Y. Danos, O. Mercer, A.C. Tags: adeno-associated viruses gene therapy gene-delivery vectors capsid cryo-EM structure AAVhu.37 research communications Source Type: research

The structure of MP-4 from Mucuna pruriens at 2.22   Å resolution
The structure of the MP-4 protein was previously determined at a resolution of 2.8   Å . Owing to the unavailability of gene-sequence information at the time, the side-chain assignment was carried out on the basis of a partial sequence available through Edman degradation, sequence homology to orthologs and electron density. The structure of MP-4 has now been determined at a higher resolution (2.22   Å ) in another space group and all of the structural inferences that were presented in the previous report of the structure were validated. In addition, the present data allowed an improved assignment of side ch...
Source: Acta Crystallographica Section F - February 1, 2020 Category: Biochemistry Authors: Jain, A. Kumar, A. Shikhi, M. Kumar, A. Nair, D.T. Salunke, D.M. Tags: Mucuna pruriens MP-4 high resolution plant proteins protease inhibitors research communications Source Type: research

Cocktailed fragment screening by X-ray crystallography of the antibacterial target undecaprenyl pyrophosphate synthase from Acinetobacter baumannii
Direct soaking of protein crystals with small-molecule fragments grouped into complementary clusters is a useful technique when assessing the potential of a new crystal system to support structure-guided drug discovery. It provides a robustness check prior to any extensive crystal screening, a double check for assay binding cutoffs and structural data for binding pockets that may or may not be picked out in assay measurements. The structural output from this technique for three novel fragment molecules identified to bind to the antibacterial target Acinetobacter baumannii undecaprenyl pyrophosphate synthase are reported, a...
Source: Acta Crystallographica Section F - January 1, 2020 Category: Biochemistry Authors: Thorpe, J.H. Wall, I.D. Sinnamon, R.H. Taylor, A.N. Stavenger, R.A. Tags: undecaprenyl pyrophosphate synthase fragment screening Acinetobacter baumannii research communications Source Type: research

The Escherichia coli RnlA – RnlB toxin – antitoxin complex: production, characterization and crystallization
The Escherichia coli rnlAB operon encodes a toxin – antitoxin module that is involved in protection against infection by bacteriophage T4. The full-length RnlA – RnlB toxin – antitoxin complex as well as the toxin RnlA were purified to homogeneity and crystallized. When the affinity tag is placed on RnlA, RnlB is largely lost during purification and the resulting crystals exclusively comprise RnlA. A homogeneous preparation of RnlA – RnlB containing stoichiometric amounts of both proteins could only be obtained using a His tag placed C-terminal to RnlB. Native mass spectrometry and SAXS indicate a 1...
Source: Acta Crystallographica Section F - January 1, 2020 Category: Biochemistry Authors: Garcia Rodriguez, G. Talavera Perez, A. Konijnenberg, A. Sobott, F. Michiels, J. Loris, R. Tags: toxin – antitoxin bacterial stress response macromolecular complex Escherichia coli research communications Source Type: research

The RING domain of mitochondrial E3 ubiquitin ligase 1 and its complex with Ube2D2: crystallization and X-ray diffraction
Mitochondrial E3 ubiquitin ligase 1 (MUL1) is located in the mitochondrial outer membrane and regulates various biological processes, including apoptosis, cell growth, mitophagy and mitochondrial dynamics. The C-terminal region of MUL1 faces the cytoplasm and contains the RING domain (MUL1-RING) where the Ub~E2 thioester binds. Unlike most RING-type E3 enzymes, MUL1-RING alone does not have an additional region that recruits a substrate protein, yet is still able to ubiquitylate the substrate, the p53 protein. Nevertheless, the exact mechanism of the ubiquitylation of p53 by MUL1-RING has not yet been elucidated. In order ...
Source: Acta Crystallographica Section F - January 1, 2020 Category: Biochemistry Authors: Lee, S.-O. Lee, C.-K. Ryu, K.-S. Chi, S.-W. Tags: mitochondrial E3 ubiquitin ligase 1 MUL1 RING domain MUL1-RING Ube2D2 ubiquitylation crystallization research communications Source Type: research