scholarly journals Cryo-EM structure provides insights into the dimer arrangement of the O-linked β-N-acetylglucosamine transferase OGT

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Richard W. Meek ◽  
James N. Blaza ◽  
Jil A. Busmann ◽  
Matthew G. Alteen ◽  
David J. Vocadlo ◽  
...  
Keyword(s):  
Repeat Unit ◽  
Structural Basis ◽  
Tetratricopeptide Repeat ◽  
Dimer Interface ◽  
X Ray ◽  
X Ray Crystallography ◽  
Protein Ligands ◽  
Repeat Units ◽  
Unit Region ◽  
Glcnac Transferase

AbstractThe O-linked β-N-acetylglucosamine modification is a core signalling mechanism, with erroneous patterns leading to cancer and neurodegeneration. Although thousands of proteins are subject to this modification, only a single essential glycosyltransferase catalyses its installation, the O-GlcNAc transferase, OGT. Previous studies have provided truncated structures of OGT through X-ray crystallography, but the full-length protein has never been observed. Here, we report a 5.3 Å cryo-EM model of OGT. We show OGT is a dimer, providing a structural basis for how some X-linked intellectual disability mutations at the interface may contribute to disease. We observe that the catalytic section of OGT abuts a 13.5 tetratricopeptide repeat unit region and find the relative positioning of these sections deviate from the previously proposed, X-ray crystallography-based model. We also note that OGT exhibits considerable heterogeneity in tetratricopeptide repeat units N-terminal to the dimer interface with repercussions for how OGT binds protein ligands and partners.

scholarly journals Structural basis for isoform-specific kinesin-1 recognition of Y-acidic cargo adaptors

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Stefano Pernigo ◽  
Magda S Chegkazi ◽  
Yan Y Yip ◽  
Conor Treacy ◽  
Giulia Glorani ◽  
...  
Keyword(s):  
Binding Sites ◽  
Adaptor Proteins ◽  
Structural Basis ◽  
Light Chains ◽  
Tetratricopeptide Repeat ◽  
X Ray ◽  
X Ray Crystallography ◽  
Specific Manner ◽  
Microtubule Motor ◽  
Partial Overlap

The light chains (KLCs) of the heterotetrameric microtubule motor kinesin-1, that bind to cargo adaptor proteins and regulate its activity, have a capacity to recognize short peptides via their tetratricopeptide repeat domains (KLCTPR). Here, using X-ray crystallography, we show how kinesin-1 recognizes a novel class of adaptor motifs that we call ‘Y-acidic’ (tyrosine flanked by acidic residues), in a KLC-isoform specific manner. Binding specificities of Y-acidic motifs (present in JIP1 and in TorsinA) to KLC1TPR are distinct from those utilized for the recognition of W-acidic motifs found in adaptors that are KLC- isoform non-selective. However, a partial overlap on their receptor binding sites implies that adaptors relying on Y-acidic and W-acidic motifs must act independently. We propose a model to explain why these two classes of motifs that bind to the concave surface of KLCTPR with similar low micromolar affinity can exhibit different capacities to promote kinesin-1 activity.

scholarly journals Structural basis for isoform-specific kinesin-1 recognition of Y-acidic cargo adaptors

2018 ◽  
Author(s):  
Stefano Pernigo ◽  
Magda Chegkazi ◽  
Yan Y. Yip ◽  
Conor Treacy ◽  
Giulia Glorani ◽  
...  
Keyword(s):  
Binding Sites ◽  
Adaptor Proteins ◽  
Structural Basis ◽  
Light Chains ◽  
Tetratricopeptide Repeat ◽  
X Ray ◽  
X Ray Crystallography ◽  
Specific Manner ◽  
Microtubule Motor ◽  
Partial Overlap

The light chains (KLCs) of the heterotetrameric microtubule motor kinesin-1, that bind to cargo adaptor proteins and regulate its activity, have a capacity to recognize short peptides via their tetratricopeptide repeat domains (KLCTPR). Here, using X-ray crystallography, we show how kinesin-1 recognizes a novel class of adaptor motifs that we call ‘Y-acidic’ (tyrosine flanked by acidic residues), in a KLC-isoform specific manner. Binding specificities of Y-acidic motifs (present in JIP1 and in TorsinA) to KLC1TPR are distinct from those utilized for the recognition of W-acidic motifs found in adaptors that are KLC-isoform non-selective. However, a partial overlap on their receptor binding sites implies that adaptors relying on Y-acidic and W-acidic motifs must act independently. We propose a model to explain why these two classes of motifs that bind to the concave surface of KLCTPR with similar low micromolar affinity can exhibit different capacities to promote kinesin-1 activity.

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

2020 ◽  
Author(s):  
Sophie M. Travis ◽  
Kevin DAmico ◽  
I-Mei Yu ◽  
Safraz Hamid ◽  
Gabriel Ramirez-Arellano ◽  
...  
Keyword(s):  
Membrane Trafficking ◽  
Relative Orientation ◽  
Snare Complex ◽  
Structural Basis ◽  
Binding Modes ◽  
Range Interaction ◽  
X Ray ◽  
X Ray Crystallography ◽  
Long Range Interaction ◽  
Target Membrane

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

scholarly journals Crystal structure of glycosyltrehalose synthase fromSulfolobus shibataeDSM5389

2018 ◽  
Vol 74 (11) ◽  
pp. 741-746
Author(s):  
Nobuo Okazaki ◽  
Michael Blaber ◽  
Ryota Kuroki ◽  
Taro Tamada
Keyword(s):  
Crystal Structure ◽  
Amino Acids ◽  
Catalytic Mechanism ◽  
Catalytic Domain ◽  
Structural Basis ◽  
Hyperthermophilic Archaeon ◽  
X Ray ◽  
X Ray Crystallography ◽  
Sulfolobus Shibatae ◽  
Glucosidic Bond

Glycosyltrehalose synthase (GTSase) converts the glucosidic bond between the last two glucose residues of amylose from an α-1,4 bond to an α-1,1 bond, generating a nonreducing glycosyl trehaloside, in the first step of the biosynthesis of trehalose. To better understand the structural basis of the catalytic mechanism, the crystal structure of GTSase from the hyperthermophilic archaeonSulfolobus shibataeDSM5389 (5389-GTSase) has been determined to 2.4 Å resolution by X-ray crystallography. The structure of 5389-GTSase can be divided into five domains. The central domain contains the (β/α)8-barrel fold that is conserved as the catalytic domain in the α-amylase family. Three invariant catalytic carboxylic amino acids in the α-amylase family are also found in GTSase at positions Asp241, Glu269 and Asp460 in the catalytic domain. The shape of the catalytic cavity and the pocket size at the bottom of the cavity correspond to the intramolecular transglycosylation mechanism proposed from previous enzymatic studies.

scholarly journals Structural Basis for Norovirus Inhibition by Human Milk Oligosaccharides

2016 ◽  
Vol 90 (9) ◽  
pp. 4843-4848 ◽  
Author(s):  
Stefan Weichert ◽  
Anna Koromyslova ◽  
Bishal K. Singh ◽  
Satoko Hansman ◽  
Stefan Jennewein ◽  
...  
Keyword(s):  
Human Milk ◽  
Blood Group ◽  
Structural Basis ◽  
Blood Group Antigens ◽  
Human Milk Oligosaccharides ◽  
Milk Oligosaccharides ◽  
X Ray ◽  
X Ray Crystallography ◽  
Naturally Occurring

Histo-blood group antigens (HBGAs) are important binding factors for norovirus infections. We show that two human milk oligosaccharides, 2′-fucosyllactose (2′FL) and 3-fucosyllactose (3FL), could block norovirus from binding to surrogate HBGA samples. We found that 2′FL and 3FL bound at the equivalent HBGA pockets on the norovirus capsid using X-ray crystallography. Our data revealed that 2′FL and 3FL structurally mimic HBGAs. These results suggest that 2′FL and 3FL might act as naturally occurring decoys in humans.

scholarly journals The search for a structural basis for therapeutic intervention against the SARS coronavirus

2007 ◽  
Vol 64 (1) ◽  
pp. 204-213 ◽  
Author(s):  
Mark Bartlam ◽  
Xiaoyu Xue ◽  
Zihe Rao
Keyword(s):  
Infectious Disease ◽  
Severe Acute Respiratory Syndrome ◽  
Structural Biology ◽  
Therapeutic Intervention ◽  
Protein Structures ◽  
Structural Basis ◽  
X Ray ◽  
X Ray Crystallography ◽  
Main Protease ◽  
Underlying Mechanisms

The 2003 outbreak of severe acute respiratory syndrome (SARS), caused by a previously unknown coronavirus called SARS-CoV, had profound social and economic impacts worldwide. Since then, structure–function studies of SARS-CoV proteins have provided a wealth of information that increases our understanding of the underlying mechanisms of SARS. While no effective therapy is currently available, considerable efforts have been made to develop vaccines and drugs to prevent SARS-CoV infection. In this review, some of the notable achievements made by SARS structural biology projects worldwide are examined and strategies for therapeutic intervention are discussed based on available SARS-CoV protein structures. To date, 12 structures have been determined by X-ray crystallography or NMR from the 28 proteins encoded by SARS-CoV. One key protein, the SARS-CoV main protease (Mpro), has been the focus of considerable structure-based drug discovery efforts. This article highlights the importance of structural biology and shows that structures for drug design can be rapidly determined in the event of an emerging infectious disease.

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