NMR spectroscopy

  • Article
    | Open Access

    Both ubiquitin and NEDD8 can be phosphorylated, but the biological role of NEDD8 phosphorylation remains unclear. Here, the authors identify similarities and differences of ubiquitin and NEDD8 phosphorylation, showing that phosphorylated NEDD8 has a distinct interactome and regulates HSP70 proteins.

    • Katrin Stuber
    • , Tobias Schneider
    •  & Martin Scheffner
  • Article
    | Open Access

    Here, the authors use solid-state NMR and EPR measurements to characterise the ATP hydrolysis transition state of the oligomeric bacterial DnaB helicase from Helicobacter pylori, which was trapped by using aluminium fluoride as a chemical mimic. They identify protein protons that coordinate to the phosphate groups of ADP and DNA and observe that the aluminium fluoride unit is highly mobile and fast-rotating.

    • Alexander A. Malär
    • , Nino Wili
    •  & Thomas Wiegand
  • Article
    | Open Access

    m6A RNA post-transcriptional modification changes RNA hybridization kinetics. Here the authors show that the methylamino group can adopt syn-conformation pairing with uridine with a mismatch-like conformation in RNA duplex. They also develop a quantitative model that predicts how m6A affects the kinetics of hybridization.

    • Bei Liu
    • , Honglue Shi
    •  & Hashim M. Al-Hashimi
  • Article
    | Open Access

    Translational regulation by riboswitches is an important mechanism for the modulation of gene expression in bacteria. Here the authors show that the ligand-induced allosteric switch in the adenine-sensing riboswitch from V. vulnificus is insufficient and leads only to a partial opening of the ribosome binding site and requires interaction with 30S-bound ribosomal protein S1, which acts as an RNA chaperone.

    • Vanessa de Jesus
    • , Nusrat S. Qureshi
    •  & Boris Fürtig
  • Article
    | Open Access

    Structural and morphological control of crystalline nanoparticles is crucial in heterogeneous catalysis. Applying DFT-assisted solid-state NMR spectroscopy, we determine the surface crystal and electronic structure of Ni2P nanoparticles, unveiling NMR nanocrystallography as an emerging tool in facet-engineered nanocatalysts.

    • Wassilios Papawassiliou
    • , José P. Carvalho
    •  & Andrew J. Pell
  • Article
    | Open Access

    The authors present a method for the conversion of full-length tau protein into seeding-competent amyloid fibrils without heparin or other negatively charged co-factors, which could be useful for studying the effects of post-translational modifications on Tau aggregation as well as to identify potential inhibitors of tau aggregation. Biochemical experiments and solid-state NMR spectroscopy measurements show that these co-factor-free tau fibrils have similar properties as amyloid fibrils isolated from brain tissue but differ from those of commonly used heparin-induced tau fibrils.

    • Pijush Chakraborty
    • , Gwladys Rivière
    •  & Markus Zweckstetter
  • Article
    | Open Access

    Tapasin is part of the peptide loading complex necessary for presenting antigenic peptides on MHC-I for the induction of adaptive immunity. Here the authors show that tapasin interacts with MHC-I in both conserved and allele-specific regions to promote antigen presentation, with tapasin L18 and K16 residues both implicated in this molecular interaction.

    • Huan Lan
    • , Esam T. Abualrous
    •  & Christian Freund
  • Article
    | Open Access

    Chromosomal translocations involving the AF10 gene, especially with CALM, are associated with aggressive leukemias. Here the authors show that the PZP domain of AF10, a histone reader, is always excluded/impaired in AF10 fusions, whereas incorporation of this domain downregulates Hoxa genes and blocks leukemogenesis.

    • Brianna J. Klein
    • , Anagha Deshpande
    •  & Tatiana G. Kutateladze
  • Article
    | Open Access

    Nuclear spin polarization and relaxation can be studied using nuclear magnetic resonance (NMR). Here the authors demonstrate a combination of fast-field cycling and optical magnetometry techniques, to realize a NMR sensor that operates in the region of very low frequency and high relaxation rate.

    • Sven Bodenstedt
    • , Morgan W. Mitchell
    •  & Michael C. D. Tayler
  • Article
    | Open Access

    The most frequent cause of familial Amyotrophic Lateral Sclerosis (ALS) and Frontotemporal Dementia (FTD) are hexanucleotide repeat expansions in the non-coding region of the C9ORF72 gene that are translated into five dipeptide repeat (DPR) proteins. Here, the authors show that proline/arginine (PR) DPRs inhibit the prolyl isomerase PPIA and reveal the molecular mechanism of the impaired protein folding activity of PPIA by performing NMR measurements and determining a PR DPR bound PPIA crystal structure.

    • Maria Babu
    • , Filippo Favretto
    •  & Markus Zweckstetter
  • Article
    | Open Access

    The molecular chaperones tapasin and TAPBPR play important roles in defining the repertoire of peptides displayed by MHC class I. Here, the authors combine NMR, ITC, fluorescence polarization measurements and deep mutational scanning analyses to reveal a peptide editing mechanism, where the G24-R36 loop in TAPBPR acts as a molecular trap to promote the selection of high-affinity peptide cargo.

    • Andrew C. McShan
    • , Christine A. Devlin
    •  & Nikolaos G. Sgourakis
  • Article
    | Open Access

    AtEH/Pan1 proteins contain two N-terminal Eps15 homology (EH) domains and are subunits of the endocytic TPLATE complex present in plants. Here, the authors combine X-ray crystallography, NMR and MD simulations with biochemical and in planta analysis to characterize the two AtEH1/Pan1 EH domains and reveal their structural differences and complementary functional roles.

    • Klaas Yperman
    • , Anna C. Papageorgiou
    •  & Daniel Van Damme
  • Article
    | Open Access

    Determining the structure of amorphous solids is important for optimization of pharmaceutical formulations, but direct relation of molecular dynamics (MD) simulations and NMR to achieve this is challenging. Here, the authors use a machine learning model of chemical shifts to solve the atomic-level structure of the hydrated amorphous drug AZD5718 by combining dynamic nuclear polarization-enhanced solid-state NMR with predicted shifts for MD simulations of large systems.

    • Manuel Cordova
    • , Martins Balodis
    •  & Lyndon Emsley
  • Article
    | Open Access

    Regulation of the MLL family of histone H3K4 methyltransferases on the nucleosome core particle (NCP) remains largely unknown. Here the authors show that intrinsically disordered regions of ASH2L and DPY30 restrict the rotational dynamics of MLL1 on the NCP, allowing more efficient enzyme-substrate engagement and higher H3K4 trimethylation activity.

    • Young-Tae Lee
    • , Alex Ayoub
    •  & Yali Dou
  • Article
    | Open Access

    The allosteric regulation of the bienzyme complex imidazole glycerol phosphate synthase (HisFH) remains to be elucidated. Here, the authors provide structural insights into the dynamic allosteric mechanism by which ligand binding to the cyclase and glutaminase active sites of HisFH regulate enzyme activation.

    • Jan Philip Wurm
    • , Sihyun Sung
    •  & Remco Sprangers
  • Article
    | Open Access

    HIV-1 integrase (IN) binds the host factor INI1/SMARCB1, which is required at multiple stages of HIV-1 replication. Here, the authors show that the same IN residues are involved in INI1 and RNA binding and in influencing particle morphogenesis and suggest that the IN-binding INI1 domain is structurally similar to HIV TAR RNA.

    • Updesh Dixit
    • , Savita Bhutoria
    •  & Ganjam V. Kalpana
  • Article
    | Open Access

    The intrinsically disordered acidic activation domain (AD) of the yeast transcription factor Gal4 acts through binding to the Med15 subunit of the Mediator complex. Here, the authors show that Gal4 interacts with Med15 through an identical fuzzy binding mechanism as Gcn4 AD, which has a different sequence, revealing a common sequence-independent mechanism for AD-Mediator binding. In contrast, Gal4 AD binds to the Gal80 repressor as a structured polypeptide, which strongly suggests that the structured binding partner dictates the type of protein–protein interaction for an intrinsically disordered protein.

    • Lisa M. Tuttle
    • , Derek Pacheco
    •  & Rachel E. Klevit
  • Article
    | Open Access

    Buried charged networks in proteins are often important for their biological functionality and are believed to destabilise the protein fold. Here, the authors combine computational design, MD simulations, biophysical experiments, NMR and X-ray crystallography to design and characterise artificial 4α-helical proteins with buried charged elements. They analyse their conformational landscapes and observe that the ion-pairs are stabilised by amphiphilic residues that electrostatically shield the charged motif, which increases structural stability.

    • Mona Baumgart
    • , Michael Röpke
    •  & Ville R. I. Kaila
  • Article
    | Open Access

    Previous work identified goddard as a putative de novo evolved gene in Drosophila melanogaster. Here, the authors characterize the structure and function of the Goddard protein in D. melanogaster, and they infer its ancestral and extant structures across the Drosophila genus.

    • Andreas Lange
    • , Prajal H. Patel
    •  & Erich Bornberg-Bauer
  • Article
    | Open Access

    Cooperation between the ULK complex and autophagy receptors mediates targeting cargoes to autophagosomes. Here, the authors show that interactions of ULK subunit FIP200 with autophagy receptors CCPG1 and Optineurin can be regulated by phosphorylation, suggesting a general binding mode shared by autophagy receptors.

    • Zixuan Zhou
    • , Jianping Liu
    •  & Lifeng Pan
  • Article
    | Open Access

    The Hsp70/Hsp40 system plays an important role in maintaining cellular proteostasis but so far it is not well understood how Hsp70 proteins are recruited to specific Hsp40 co-chaperones. Here, the authors combine biochemical and biophysical approaches to characterise the oligomeric mammalian Hsp40 DnaJB8. They identify an intra-oligomer DnaJB8 interaction between the N-terminal J-Domain and the C-terminal domain that occludes the J-Domain surface that binds Hsp70 and propose a model for DnaJB8-Hsp70 recruitment.

    • Bryan D. Ryder
    • , Irina Matlahov
    •  & Lukasz A. Joachimiak
  • Article
    | Open Access

    α-Synuclein is a presynaptic protein whose aberrant aggregation is associated with Parkinson’s disease. Here, the authors show how αSynuclein-induced docking of synaptic vesicles is modulated by the lipid composition changes typically observed in neurodegeneration using an in vitro system.

    • Wing K. Man
    • , Bogachan Tahirbegi
    •  & Giuliana Fusco
  • Article
    | Open Access

    Spy is an ATP independent chaperone that can act as both a holdase and a foldase towards topologically simple substrates. Assessing the interaction of Spy and apoflavodoxin, a complex client, the authors show that Spy’s activity is substrate specific. Spy binds partially unfolded states of apoflavodoxin tightly, which limits the possibility of folding and converts Spy to a pure holdase.

    • Rishav Mitra
    • , Varun V. Gadkari
    •  & James C. A. Bardwell
  • Article
    | Open Access

    p23 is a co-chaperone of Hsp90 but its mode of action is mechanistically not well understood. Here, the authors combine in vitro and yeast in vivo assays, biochemical measurements and NMR experiments to characterize p23 and identify two conserved helical elements in the intrinsically disordered C-terminal tail of p23 that together with the folded domain of p23 regulate the Hsp90 ATPase activity and affect the binding and maturation of Hsp90 clients.

    • Maximilian M. Biebl
    • , Abraham Lopez
    •  & Johannes Buchner
  • Article
    | Open Access

    Here, the authors present Methyl Assignments Using Satisfiability (MAUS), a method for the assignment of methyl groups using raw NOE data. They use eight proteins in the 10–45 kDa size range as test cases and show that MAUS yields 100% accurate assignments at high completeness levels.

    • Santrupti Nerli
    • , Viviane S. De Paula
    •  & Nikolaos G. Sgourakis
  • Article
    | Open Access

    The E2-like enzyme human Atg3 catalyses the transfer of ubiquitin-like mammalian LC3 to the lipid phosphatidylethanolamine during autophagosome formation. Here, the authors combine NMR measurements with in vitro biochemical and in vivo cellular assays and show that the N-terminal conserved region of human Atg3 communicates information from the curvature-sensing domain to its active site.

    • Yansheng Ye
    • , Erin R. Tyndall
    •  & Fang Tian
  • Article
    | Open Access

    The small proton-coupled transporter EmrE confers multidrug resistance in bacteria. The structure of drug-bound EmrE in phospholipid bilayers is now determined using solid-state NMR. The structure provides detailed insights into the molecular mechanism of substrate recognition by this transporter.

    • Alexander A. Shcherbakov
    • , Grant Hisao
    •  & Mei Hong
  • Article
    | Open Access

    The authors present a method for calculating the accuracy of an NMR structure, where flexibility from backbone chemical shifts is compared to structural flexibility predicted using rigidity theory. The authors validate their method and use it to compare the accuracy of NMR and X-ray structures.

    • Nicholas J. Fowler
    • , Adnan Sljoka
    •  & Mike P. Williamson
  • Article
    | Open Access

    α1-Antitrypsin (AAT) is a 52 kDa serum glycoprotein, the misfolding and polymerisation of which is associated with COPD and liver disease. Here the authors demonstrate the use of high-resolution multidimensional solution-state NMR spectroscopy to characterise the structure and dynamics in solution of Z AAT purified directly from clinical patients.

    • Alistair M. Jagger
    • , Christopher A. Waudby
    •  & David A. Lomas
  • Article
    | Open Access

    Cyclophilin A (CypA) is a peptidylprolyl isomerase that also has chaperone activity and interacts with the intrinsically disordered protein α-Synuclein (aSyn). Here, the authors combine NMR measurements and biochemical experiments to characterise the interplay between the catalysis of proline isomerization and molecular chaperone activity of CypA and find that both activities have opposing effects on aSyn and further show that the that cis/trans isomerization outpowers the holding activity of CypA.

    • Filippo Favretto
    • , David Flores
    •  & Markus Zweckstetter
  • Article
    | Open Access

    Sorghum is a source of lignocellulosic biomass for the production of renewable fuels. Here the authors characterise the sorghum secondary cell wall using multi-dimensional magic angle spinning solid-state NMR and present a model dominated by interactions between three-fold screw xylan and amorphous cellulose.

    • Yu Gao
    • , Andrew S. Lipton
    •  & Jenny C. Mortimer
  • Article
    | Open Access

    The SARS-CoV-2 viral genome is encapsulated by the nucleocapsid protein (NSARS-CoV-2) that is essential for viral replication. Here, the authors show that RNA induces liquid-liquid phase separation of NSARS-CoV-2 and how NSARS-CoV-2 phosphorylation modulates RNA-binding and phase separation and that these RNA/NSARS-CoV-2-droplets recruit and concentrate the SARS-CoV-2 RNA-dependent RNA polymerase complex in vitro, which would enable high initiation and elongation rates during viral transcription.

    • Adriana Savastano
    • , Alain Ibáñez de Opakua
    •  & Markus Zweckstetter
  • Article
    | Open Access

    Here, the authors combine 3D printing and liquid metal filling techniques to fabricate customised probeheads for magnetic resonance experiments. They demonstrate in situ electrochemical nuclear magnetic resonance analysis, reaction monitoring with continues-flow separation and small-sample imaging.

    • Junyao Xie
    • , Xueqiu You
    •  & Zhong Chen
  • Article
    | Open Access

    Bacteriophages of the Siphoviridae family have a long, flexible, non-contractile tail that has been difficult to characterize structurally. Here, the authors present the atomic structure of the tail tube of one of these phages, showing a hollow flexible tube formed by hexameric rings stacked by flexible linkers.

    • Maximilian Zinke
    • , Katrin A. A. Sachowsky
    •  & Adam Lange
  • Article
    | Open Access

    The low-complexity (LC) domain mediates liquid-liquid phase separation and fibril formation of the RNA-binding protein FUS (FUsed in Sarcoma). Here, the authors combine cryo-EM, solid-state NMR measurements and MD simulations to structurally characterise the fibrils formed by the C-terminal half of the FUS LC domain and discuss stabilizing interactions within the fibril core.

    • Myungwoon Lee
    • , Ujjayini Ghosh
    •  & Robert Tycko
  • Article
    | Open Access

    So far only a few compounds have been reported as splicing modulators. Here, the authors combine high-throughput screening, chemical synthesis, NMR, X-ray crystallography with functional studies and develop phenothiazines as inhibitors for the U2AF Homology Motif (UHM) domains of proteins that regulate splicing and show that they inhibit early spliceosome assembly on pre-mRNA substrates in vitro.

    • Pravin Kumar Ankush Jagtap
    • , Tomáš Kubelka
    •  & Michael Sattler
  • Article
    | Open Access

    As protein synthesis takes place, newly synthesized polypeptide chain passes through the ribosomal exit tunnel, which can accommodate up to 70 residues in the case of a helical peptide. Here the authors show that oxidation of cysteine residues in the nascent chain can occur within the ribosome exit tunnel, where sufficient space exists for the formation of disulfide bonds.

    • Linda Schulte
    • , Jiafei Mao
    •  & Harald Schwalbe
  • Article
    | Open Access

    The structure and dynamics of large proteins and complexes can be studied by methyl-NMR but resonance assignment is still challenging. Here, the authors present a NMR method that leverages optimal control pulse design to unambiguously distinguish between Leu and Val using a simple 2D HMQC experiment and they apply it to several proteins including Cas9, interleukin, and human translation initiation factor eIF4a.

    • Soumya P. Behera
    • , Abhinav Dubey
    •  & Haribabu Arthanari