TY  - JOUR
AB  - Incorporating metal cofactors into computationally designed protein scaffolds provides a versatile route to novel protein functions, including the potential for new-to-nature enzyme catalysis. However, a major challenge in protein design is to understand how the scaffold architecture influences conformational dynamics. Here, we characterized structure and dynamics of a modular de novo scaffold with flexible inter-domain linkers. Three rationally engineered variants with different metal specificity were studied by combining X-ray crystallography, NMR spectroscopy, and molecular dynamics simulations. The lanthanide-binding variant was initially trapped in an inactive conformational state, which impaired efficient metal coordination and cerium-dependent photocatalytic activity. Stabilization of the active conformation by AI-guided sequence optimization using ProteinMPNN led to accelerated lanthanide binding and a 10-fold increase in kcat/Km for a photoenzymatic model reaction. Our results suggest that modular scaffold architectures provide an attractive starting point for de novo metalloenzyme engineering and that ProteinMPNN-based sequence redesign can stabilize desired conformational states.
AU  - Wagner Egea, P.*
AU  - Delhommel, F.
AU  - Mustafa, G.*
AU  - Leiss-Maier, F.*
AU  - Klimper, L.*
AU  - Badmann, T.*
AU  - Heider, A.*
AU  - Wille, I.*
AU  - Groll, M.*
AU  - Sattler, M.
AU  - Zeymer, C.*
C1  - 75977
C2  - 58299
TI  - Modular protein scaffold architecture and AI-guided sequence optimization facilitate de novo metalloenzyme engineering.
JO  - Structure
PY  - 2025
SN  - 0969-2126
ER  - 

TY  - CONF
AB  - Conformational dynamics is crucial for the biological function of RNA molecules and for their potential as therapeutic targets. This meeting report outlines key "take-home" messages that emerged from the presentations and discussions during the CECAM workshop "RNA dynamics from experimental and computational approaches" in Paris, June 26-28, 2023.
AU  - Bussi, G.*
AU  - Bonomi, M.*
AU  - Gkeka, P.*
AU  - Sattler, M.
AU  - Al-Hashimi, H.M.*
AU  - Auffinger, P.*
AU  - Duca, M.*
AU  - Foricher, Y.*
AU  - Incarnato, D.*
AU  - Jones, A.N.*
AU  - Kirmizialtin, S.*
AU  - Krepl, M.*
AU  - Orozco, M.*
AU  - Palermo, G.*
AU  - Pasquali, S.*
AU  - Salmon, L.*
AU  - Schwalbe, H.*
AU  - Westhof, E.*
AU  - Zacharias, M.*
C1  - 71643
C2  - 56329
SP  - 1281-1287
TI  - RNA dynamics from experimental and computational approaches.
JO  - Structure
VL  - 32
IS  - 9
PY  - 2024
SN  - 0969-2126
ER  - 

TY  - JOUR
AB  - During RNA replication, coronaviruses require proofreading to maintain the integrity of their large genomes. Nsp14 associates with viral polymerase complex to excise the mismatched nucleotides. Aside from the exonuclease activity, nsp14 methyltransferase domain mediates cap methylation, facilitating translation initiation and protecting viral RNA from recognition by the innate immune sensors. The nsp14 exonuclease activity is modulated by a protein co-factor nsp10. While the nsp10/nsp14 complex structure is available, the mechanistic basis for nsp10-mediated modulation remains unclear in the absence of the nsp14 structure. Here, we provide a crystal structure of nsp14 in an apo-form. Comparative analysis of the apo- and nsp10-bound structures explain the modulatory role of the co-factor protein and reveal the allosteric nsp14 control mechanism essential for drug discovery. Further, the flexibility of the N-terminal lid of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nsp14 structure presented in this study rationalizes the recently proposed idea of nsp14/nsp10/nsp16 ternary complex.
AU  - Czarna, A.*
AU  - Plewka, J.*
AU  - Kresik, L.*
AU  - Matsuda, A.*
AU  - Karim, A.*
AU  - Robinson, C.*
AU  - O'Byrne, S.*
AU  - Cunningham, F.*
AU  - Georgiou, I.*
AU  - Wilk, P.*
AU  - Pachota, M.*
AU  - Popowicz, G.M.
AU  - Wyatt, P.G.*
AU  - Dubin, G.*
AU  - Pyrc, K.*
C1  - 65064
C2  - 52654
SP  - 1050-1054.e2
TI  - Refolding of lid subdomain of SARS-CoV-2 nsp14 upon nsp10 interaction releases exonuclease activity.
JO  - Structure
VL  - 30
IS  - 8
PY  - 2022
SN  - 0969-2126
ER  - 

TY  - JOUR
AB  - In this issue of Structure, Urbanek et al. (2020a) combine site-specific isotope labeling and NMR spectroscopy to investigate opposing effects of flanking regions onto the conformation of the poly-Q region in Huntingtin. Poly-Q interactions with preceding residues promote an a-helical conformation while a following proline-rich region favors extended conformations.
AU  - Delhommel, F.
AU  - Sattler, M.
C1  - 59573
C2  - 48920
CY  - 50 Hampshire St, Floor 5, Cambridge, Ma 02139 Usa
SP  - 730-732
TI  - When less is more: Combining site-specific isotope labeling and NMR unravels structural details of huntingtin repeats.
JO  - Structure
VL  - 28
IS  - 7
PB  - Cell Press
PY  - 2020
SN  - 0969-2126
ER  - 

TY  - JOUR
AB  - Pdx1 is a transcription factor crucial for development and maintenance of a functional pancreas. It regulates insulin expression and glucose homeostasis. SPOP is an E3-ubiquitin ligase adaptor protein that binds Pdx1, thus triggering its ubiquitination and proteasomal degradation. However, the underlying mechanisms are not well understood. Here, we present the crystal structure of the SPOP-Pdx1 complex. We show that Pdx1 residues 223-233 bind to SPOP MATH domain with low micromolar affinity. The interface is extended compared to other SPOP-client proteins. Previously, Pdx1 phosphorylation has been proposed to have a regulatory function. In this respect we show that phosphorylation lowers the affinity of Pdx1 to SPOP by isothermal titration calorimetry and nuclear magnetic resonance data. Our data provide insights into a critical protein-protein interaction that regulates cellular Pdx1 levels by SPOP-mediated decay. A reduction of Pdx1 levels in beta cells is linked to apoptosis and considered a hallmark of type 2 diabetes.
AU  - Ostertag, M.S.
AU  - Messias, A.C.
AU  - Sattler, M.
AU  - Popowicz, G.M.
C1  - 54777
C2  - 45887
CY  - 50 Hampshire St, Floor 5, Cambridge, Ma 02139 Usa
SP  - 327-334.e3
TI  - The structure of the SPOP-Pdx1 interface reveals insights into the phosphorylation-dependent binding regulation.
JO  - Structure
VL  - 27
IS  - 2
PB  - Cell Press
PY  - 2019
SN  - 0969-2126
ER  - 

TY  - JOUR
AB  - Ric-8A is a 530-amino acid cytoplasmic molecular chaperone and guanine nucleotide exchange factor (GEF) for i, q, and 12/13 classes of heterortrimeric G protein alpha subunits (G alpha). We report the 2.2-angstrom crystal structure of the Ric-8A G alpha-binding domain with GEF activity, residues 1-452, and is phosphorylated at Ser435 and Thr440. Residues 1-429 adopt a superhelical fold comprised of Armadillo (ARM) and HEAT repeats, and the C terminus is disordered. One of the phosphorylated residues potentially binds to a basic cluster in an ARM motif. Amino acid sequence conservation and published hydrogen- deuterium exchange data indicate repeats 3 through 6 to be a putative G alpha-binding surface. Normal mode modeling of small-angle X-ray scattering data indicates that phosphorylation induces relative rotation between repeats 1-4, 5-6, and 7-9. 2D H-1-N-1(5)-TROSY spectra of [H-2, N-15]-labeled Gail in the presence of R452 reveals chemical shift perturbations of the C terminus and Gail residues involved in nucleotide binding.
AU  - Zeng, B.*
AU  - Mou, T.C.*
AU  - Doukov, T.I.*
AU  - Steiner, A.
AU  - Yu, W.*
AU  - Papasergi-Scott, M.*
AU  - Tall, G.G.*
AU  - Hagn, F.
AU  - Sprang, S.R.*
C1  - 56202
C2  - 46896
CY  - 50 Hampshire St, Floor 5, Cambridge, Ma 02139 Usa
SP  - 1137-1147.e5
TI  - Structure, function, and dynamics of the G alpha binding domain of Ric-8A.
JO  - Structure
VL  - 27
IS  - 7
PB  - Cell Press
PY  - 2019
SN  - 0969-2126
ER  - 

TY  - JOUR
AB  - Reactivation of p53 by release of the functional protein from its inhibition by MDM2 provides an efficient, nongenotoxic approach to a wide variety of cancers. We present the cocrystal structures of two complexes of MDM2 with inhibitors based on 6-chloroindole scaffolds. Both molecules bound to a distinct conformational state of MDM2 with nM-μM affinities. In contrast to other structurally characterized antagonists, which mimic three amino acids of p53 (Phe19, Trp23, and Leu26), the compounds induced an additional hydrophobic pocket on the MDM2 surface and unveiled a four-point binding mode. The enlarged interaction interface of the inhibitors resulted in extension of small molecules binding toward the "lid" segment of MDM2 (residues 19-23)-a nascent element that interferes with p53 binding. As supported by protein engineering and molecular dynamics studies, employing these unstable elements of MDM2 provides an efficient and yet unexplored alternative in development of MDM2-p53 association inhibitors.
AU  - Bista, M.*
AU  - Wolf, S.*
AU  - Khoury, K.*
AU  - Kowalska, K.*
AU  - Huang, Y.*
AU  - Wrona, E.*
AU  - Arciniega, M.*
AU  - Popowicz, G.M.*
AU  - Holak, T.A.*
AU  - Dömling, A.*
C1  - 28098
C2  - 32932
SP  - 2143-2151
TI  - Transient protein states in designing inhibitors of the MDM2-p53 interaction.
JO  - Structure
VL  - 21
IS  - 12
PB  - Cell Press
PY  - 2013
SN  - 0969-2126
ER  - 

TY  - JOUR
AB  - ADP-ribosyltransferases (ARTs) catalyze the transfer of ADP-ribose from NAD(+) onto substrates. Some ARTs generate in an iterative process ADP-ribose polymers that serve as adaptors for distinct protein domains. Other ARTs, exemplified by ARTD10, function as mono-ADP-ribosyltransferases, but it has been unclear whether this modification occurs in cells and how it is read. We observed that ARTD10 colocalized with ARTD8 and defined its macrodomains 2 and 3 as readers of mono-ADP-ribosylation both in vitro and in cells. The crystal structures of these two ARTD8 macrodomains and isothermal titration calorimetry confirmed their interaction with ADP-ribose. These macrodomains recognized mono-ADP-ribosylated ARTD10, but not poly-ADP-ribosylated ARTD1. This distinguished them from the macrodomain of macroH2A1.1, which interacted with poly- but not mono-ADP-ribosylated substrates. Moreover, Ran, an ARTD10 substrate, was also read by ARTD8 macrodomains. This identifies readers of mono-ADP-ribosylated proteins, defines their structures, and demonstrates the presence of this modification in cells.
AU  - Forst, A.H.*
AU  - Karlberg, T.*
AU  - Herzog, N.*
AU  - Thorsell, A.-G.*
AU  - Gross, A.*
AU  - Feijs, K.L.H.*
AU  - Verheugd, P.*
AU  - Kursula, P.*
AU  - Nijmeijer, B.*
AU  - Kremmer, E.
AU  - Kleine, H.*
AU  - Ladurner, A.G.*
AU  - Schüler, H.*
AU  - Lüscher, B.*
C1  - 23635
C2  - 31271
SP  - 462-475
TI  - Recognition of mono-ADP-ribosylated ARTD10 substrates by ARTD8 macrodomains.
JO  - Structure
VL  - 21
IS  - 3
PB  - Cell Press
PY  - 2013
SN  - 0969-2126
ER  - 

TY  - JOUR
AB  - CUG-binding protein 1 (CUGBP1) is a ubiquitous RNA-binding protein implicated in altered RNA metabolism linked to myotonic dystrophy type 1. Crystal structures of the RRM domains in complex with cognate RNAs (Teplova et al., 2010) reveal molecular details for the selectivity of CUGBP1 toward GU-rich mRNA elements.
AU  - Tripsianes, K.
AU  - Sattler, M.
C1  - 3834
C2  - 28366
SP  - 1228-1229
TI  - Repeat recognition.
JO  - Structure
VL  - 18
IS  - 10
PY  - 2010
SN  - 0969-2126
ER  - 

TY  - JOUR
AB  - The localization of mRNAs in subcellular compartments is an efficient way to spatially restrict gene expression. Crystal structures of raver1-vinculin reported by Izard and coworkers now suggest a possible mechanism for mRNA localization during the assembly of focal adhesions.
AU  - Madl, T.
AU  - Sattler, M.
C1  - 257
C2  - 26886
SP  - 781-783
TI  - Adhesion dance with raver.
JO  - Structure
VL  - 17
IS  - 6
PB  - Cell Press
PY  - 2009
SN  - 0969-2126
ER  -