Structural basis for methyl-donor–dependent and sequence-specific binding to tRNA substrates by knotted methyltransferase TrmD

The deep trefoil knot architecture is unique to the SpoU and tRNA methyltransferase D (TrmD) (SPOUT) family of methyltransferases (MTases) in all three domains of life. In bacteria, TrmD catalyzes the N1-methylguanosine (m1G) modification at position 37 in transfer RNAs (tRNAs) with the 36GG37 sequence, using S-adenosyl-l-methionine (AdoMet) as the methyl donor. The m1G37-modified tRNA functions properly to prevent +1 frameshift errors on the ribosome. Here we report the crystal structure of the TrmD homodimer in complex with a substrate tRNA and an AdoMet analog. Our structural analysis revealed the mechanism by which TrmD binds the substrate tRNA in an AdoMet-dependent manner. The trefoil-knot center, which is structurally conserved among SPOUT MTases, accommodates the adenosine moiety of AdoMet by loosening/retightening of the knot. The TrmD-specific regions surrounding the trefoil knot recognize the methionine moiety of AdoMet, and thereby establish the entire TrmD structure for global interactions with tRNA and sequential and specific accommodations of G37 and G36, resulting in the synthesis of m1G37-tRNA.

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Functional assignment based on structural analysis: Crystal structure of the yggJ protein (HI0303) ofHaemophilus influenzae reveals an RNA methyltransferase with a deep trefoil knot

Proteins Structure Function and Bioinformatics ◽

10.1002/prot.10510 ◽

2003 ◽

Vol 53 (2) ◽

pp. 329-332 ◽

Cited By ~ 21

Author(s):

Farhad Forouhar ◽

Jianwei Shen ◽

Rong Xiao ◽

Thomas B. Acton ◽

Gaetano T. Montelione ◽

...

Keyword(s):

Crystal Structure ◽

Structural Analysis ◽

Functional Assignment ◽

Trefoil Knot ◽

Ofhaemophilus Influenzae

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The crystal structure of TRPM2 MHR1/2 domain reveals a conserved Zn2+-binding domain essential for ligand binding and activity

10.1101/2022.01.04.474898 ◽

2022 ◽

Author(s):

Simon Sander ◽

Ellen Gattkowski ◽

Jelena Pick ◽

Ralf Fliegert ◽

Henning Tidow

Keyword(s):

Crystal Structure ◽

Transient Receptor Potential ◽

Specific Binding ◽

Receptor Potential ◽

Binding Domain ◽

Dependent Manner ◽

Structural Element ◽

Transient Receptor Potential Melastatin ◽

Transient Receptor ◽

Species Specific

Transient receptor potential melastatin 2 (TRPM2) is a Ca2+-permeable, non-selective cation channel involved in diverse physiological processes such as immune response, apoptosis and body temperature sensing. TRPM2 is activated by ADP-ribose (ADPR) and 2′-deoxy-ADPR in a Ca2+-dependent manner. While two species-specific binding sites exist for ADPR, a binding site for 2′-deoxy-ADPR is not known yet. Here, we report the crystal structure of the MHR1/2 domain of TRPM2 from zebrafish (Danio rerio) and show binding of both ligands to this domain. We identified a so-far unrecognized Zn2+-binding domain that was not resolved in previous cryo-EM structures and that is conserved in most TRPM channels. In combination with patch clamp experiments, we comprehensively characterize the effect of the Zn2+-binding domain on TRPM2 activation. Our results provide insight into a conserved structural element essential for channel activity.

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Structural basis of human 5,10-methylenetetrahydrofolate reductase (MTHFR) regulation by phosphorylation and S-adenosylmethionine inhibition

10.1101/271593 ◽

2018 ◽

Author(s):

D. Sean Froese ◽

Jola Kopec ◽

Elzbieta Rembeza ◽

Gustavo Arruda Bezerra ◽

Anselm Erich Oberholzer ◽

...

Keyword(s):

Crystal Structure ◽

Amino Acid ◽

Methylenetetrahydrofolate Reductase ◽

Binding Domain ◽

Structural Basis ◽

Methyl Donor ◽

Tim Barrel ◽

Conformational Plasticity ◽

Methionine Cycle ◽

Regulatory Connection

AbstractThe folate and methionine cycles are crucial to the biosynthesis of lipids, nucleotides and proteins, and production of the global methyl donor S-adenosylmethionine (SAM). 5,10-methylenetetrahydrofolate reductase (MTHFR) represents a key regulatory connection between these cycles, generating 5-methyltetrahydrofolate for initiation of the methionine cycle, and undergoing allosteric inhibition by its end product SAM. Our 2.5 Å resolution crystal structure of human MTHFR reveals a unique architecture, appending the well-conserved catalytic TIM-barrel to a eukaryote-only SAM-binding domain. The latter domain of novel fold provides the predominant interface for MTHFR homo-dimerization, positioning the N-terminal serine-rich phosphorylation region into proximity with the C-terminal SAM-binding domain. This explains how MTHFR phosphorylation, identified on 11 N-terminal residues (16-total), increases sensitivity to SAM binding and inhibition. Finally, we demonstrate the 25-amino-acid inter-domain linker enables conformational plasticity and propose it to be a key mediator of SAM regulation.

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Structural basis for a natural circular permutation in proteins

10.1101/2020.10.28.360099 ◽

2020 ◽

Author(s):

Samuel G. Nonis ◽

Joel Haywood ◽

Jason W. Schmidberger ◽

Charles S. Bond ◽

Joshua S. Mylne

Keyword(s):

Crystal Structure ◽

Amino Acid ◽

Structural Analysis ◽

Circular Permutation ◽

Structural Basis ◽

Carbohydrate Binding ◽

Post Translational Modification ◽

Jack Bean ◽

Asparaginyl Endopeptidase

AbstractOver 30 years ago, an intriguing post-translational modification was discovered to be responsible for creating concanavalin A (conA), a carbohydrate-binding protein found in the seeds of jack bean (Canavalia ensiformis) and commercially used for carbohydrate chromatography. Biosynthesis of conA involves what was then an unprecedented rearrangement in amino acid sequence, whereby the N-terminal half of the gene-encoded conA precursor is swapped to become the C-terminal half of conA. The cysteine protease, asparaginyl endopeptidase (AEP), was shown to be involved, but its mechanism was not fully elucidated. To understand the structural basis and consequences of conA circular permutation, we generated a recombinant jack bean conA precursor (pro-conA) plus jack bean AEP (CeAEP1) and solved crystal structures for each to 2.1 Å and 2.7 Å respectively. By reconstituting the biosynthesis of conA in vitro, we prove CeAEP1 alone can perform both the cleavage and cleavage-coupled transpeptidation to form conA. CeAEP1 structural analysis reveals how it is capable of carrying out both these reactions. Biophysical assays illustrated that conA is more thermally and pH stable than pro-conA, consistent with fewer intermolecular interactions between subunits in the pro-conA crystal structure. These findings elucidate the consequences of circular permutation in the only post-translation example known to occur in nature.

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Structure of the complex of phosphorylated liver kinase B1 and 14-3-3ζ

Acta Crystallographica Section F Structural Biology Communications ◽

10.1107/s2053230x17003521 ◽

2017 ◽

Vol 73 (4) ◽

pp. 196-201 ◽

Cited By ~ 2

Author(s):

Yongjian Lu ◽

Sheng Ding ◽

Ruiqing Zhou ◽

Jianyong Wu

Keyword(s):

Crystal Structure ◽

Protein Kinase ◽

Fusion Protein ◽

Binding Mode ◽

Tumour Suppressor ◽

Structural Basis ◽

Dependent Manner ◽

Multiple Cancers ◽

Physiological Processes ◽

Liver Kinase B1

The serine/threonine protein kinase liver kinase B1 (LKB1) is a tumour suppressor and plays important roles in development and metabolism. It phosphorylates AMPK and AMPK-related kinases to regulate multiple physiological processes. Mutations in LKB1 often occur in multiple cancers. LKB1 can be suppressed by 14-3-3 proteins in a phosphorylation-dependent manner. Previously, the structure of a 14-3-3ζ–LKB1 fusion protein has been reported, revealing a phosphorylation-independent binding mode of LKB1 to 14-3-3 proteins. Here, the crystal structure of phosphorylated LKB1 peptide in complex with 14-3-3ζ was solved, which provides a structural basis for the phosphorylation-dependent recognition of LKB1 by 14-3-3 proteins.

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