ORCID Profile
0000-0002-7597-2133
Current Organisations
Monash University
,
Stanford University
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Cellular Immunology | Biochemistry and Cell Biology | Structural Biology (incl. Macromolecular Modelling) | Innate Immunity
Expanding Knowledge in the Biological Sciences | Immune System and Allergy |
Publisher: Elsevier BV
Date: 10-2020
Publisher: Proceedings of the National Academy of Sciences
Date: 21-09-2020
Abstract: A newly discovered system for immunological detection of erse bacterial and fungal pathogens involves the MHC-like host protein called MR1. This molecule scavenges metabolites from the biosynthesis of riboflavin by microbes. MR1 presents these compounds on the surface of antigen-presenting cells, where they interact with T cells known as mucosal-associated invariant T cells and stimulate immunity. Critical aspects of the cell biology of metabolite presentation by MR1 are unknown. Here we generated a fluorescent antigen analog and use it to show that MR1 captures its metabolites within the endoplasmic reticulum. We describe proteins that maintain MR1 ready for metabolite binding in the endoplasmic reticulum to promote efficient pathogen detection. MR1 thus monitors extracellular microbial metabolites from within the cell.
Publisher: Frontiers Media SA
Date: 27-08-2020
Publisher: Elsevier BV
Date: 09-2015
Publisher: American Chemical Society (ACS)
Date: 05-02-2018
DOI: 10.1021/ACS.BIOCHEM.7B00557
Abstract: Heparan sulfate proteoglycans are proteins substituted with one or more heparan sulfate (HS) polysaccharides, found in abundance at cell surfaces. HS chains influence the activity of many biologically important molecules involved in cellular communication and signaling. The exostosin (EXT) proteins are glycosyltransferases in the Golgi apparatus that assemble HS chains on HSPGs. The EXTL3 enzyme mainly works as an initiator in HS biosynthesis. In this work, human lumenal N-glycosylated EXTL3 (EXTL3ΔN) was cloned, expressed in human embryonic kidney cells, and purified. Various biophysical and biochemical approaches were then employed to elucidate the N-glycosylation sites and the function of their attached N-glycans. Furthermore, the stability and conformation of the purified EXTL3ΔN protein in solution have been analyzed. Our data show that EXTL3ΔN has N-glycans at least at two positions, Asn290 and Asn592, which seem to be critical for proper protein folding and/or release. EXTL3ΔN is quite stable, as high temperature (∼59 °C) was required for denaturation. Deconvolution of the EXTL3ΔN far-UV CD spectrum revealed a substantial fraction of β sheets (25%) with a minor proportion of α-helices (14%) in the secondary structure. Solution small-angle X-ray scattering and dynamic light scattering revealed an extended structure suggestive of a dimeric arrangement and consisting of two distinct regions, narrow and broad, respectively. This is consistent with bioinformatics analyses suggesting a 3-domain structure with two glycosyltransferase domains and a coiled-coil domain.
Publisher: Proceedings of the National Academy of Sciences
Date: 27-04-2020
Abstract: The antigen-presenting molecule MR1 presents riboflavin-based metabolites to Mucosal-Associated Invariant T (MAIT) cells. While MR1 egress to the cell surface is ligand-dependent, the ability of small-molecule ligands to impact on MR1 cellular trafficking remains unknown. Arising from an in silico screen of the MR1 ligand-binding pocket, we identify one ligand, 3-([2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-yl]formamido)propanoic acid, DB28, as well as an analog, methyl 3-([2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-yl]formamido)propanoate, NV18.1, that down-regulate MR1 from the cell surface and retain MR1 molecules in the endoplasmic reticulum (ER) in an immature form. DB28 and NV18.1 compete with the known MR1 ligands, 5-OP-RU and acetyl-6-FP, for MR1 binding and inhibit MR1-dependent MAIT cell activation. Crystal structures of the MAIT T cell receptor (TCR) complexed with MR1-DB28 and MR1-NV18.1, show that these two ligands reside within the A′-pocket of MR1. Neither ligand forms a Schiff base with MR1 molecules both are nevertheless sequestered by a network of hydrophobic and polar contacts. Accordingly, we define a class of compounds that inhibits MR1 cellular trafficking.
Publisher: Rockefeller University Press
Date: 26-08-2022
DOI: 10.1084/JEM.20210828
Abstract: Mucosal-associated invariant T (MAIT) cells detect microbial infection via recognition of riboflavin-based antigens presented by the major histocompatibility complex class I (MHC-I)–related protein 1 (MR1). Most MAIT cells in human peripheral blood express CD8αα or CD8αβ coreceptors, and the binding site for CD8 on MHC-I molecules is relatively conserved in MR1. Yet, there is no direct evidence of CD8 interacting with MR1 or the functional consequences thereof. Similarly, the role of CD8αα in lymphocyte function remains ill-defined. Here, using newly developed MR1 tetramers, mutated at the CD8 binding site, and by determining the crystal structure of MR1–CD8αα, we show that CD8 engaged MR1, analogous to how it engages MHC-I molecules. CD8αα and CD8αβ enhanced MR1 binding and cytokine production by MAIT cells. Moreover, the CD8–MR1 interaction was critical for the recognition of folate-derived antigens by other MR1-reactive T cells. Together, our findings suggest that both CD8αα and CD8αβ act as functional coreceptors for MAIT and other MR1-reactive T cells.
Publisher: Wiley
Date: 27-02-2018
DOI: 10.1111/IMCB.12017
Abstract: The major histocompatibility complex (MHC) class-I related molecule MR1 is a monomorphic and evolutionary conserved antigen (Ag)-presenting molecule that shares the overall architecture of MHC-I and CD1 proteins. However, in contrast to MHC-I and the CD1 family that present peptides and lipids, respectively, MR1 specifically presents small organic molecules. During microbial infection of mammalian cells, MR1 captures and presents vitamin B precursors, derived from the microbial biosynthesis of riboflavin, on the surface of antigen-presenting cells. These MR1-Ag complexes are recognized by the mucosal-associated invariant T cell receptor (MAIT TCR), which subsequently leads to MAIT cell activation. Recently, MR1 was shown to trap chemical scaffolds including drug and drug-like molecules. Here, we review this metabolite Ag-presenting molecule and further define the key molecular interactions underlying the recognition and reactivity of MAIT TCRs to MR1 in an Ag-dependent manner.
Publisher: Springer Science and Business Media LLC
Date: 02-03-2020
Publisher: Elsevier BV
Date: 04-2012
Publisher: Academic Journals
Date: 02-12-2011
DOI: 10.5897/IJPS11.1005
Publisher: International Union of Crystallography (IUCr)
Date: 19-11-2013
DOI: 10.1107/S0907444913025250
Abstract: The use of controlled dehydration for improvement of protein crystal diffraction quality is increasing in popularity, although there are still relatively few documented ex les of success. A study has been carried out to establish whether controlled dehydration could be used to improve the anisotropy of crystals of the core protein of the human proteoglycan glypican-1. Crystals were subjected to controlled dehydration using the HC1 device. The optimal protocol for dehydration was developed by careful investigation of the following parameters: dehydration rate, final relative humidity and total incubation time T inc . Of these, the most important was shown to be T inc . After dehydration using the optimal protocol the crystals showed significantly reduced anisotropy and improved electron density, allowing the building of previously disordered parts of the structure.
Publisher: Wiley
Date: 10-10-2021
DOI: 10.1002/PROT.26247
Abstract: The biological and functional significance of selected Critical Assessment of Techniques for Protein Structure Prediction 14 (CASP14) targets are described by the authors of the structures. The authors highlight the most relevant features of the target proteins and discuss how well these features were reproduced in the respective submitted predictions. The overall ability to predict three‐dimensional structures of proteins has improved remarkably in CASP14, and many difficult targets were modeled with impressive accuracy. For the first time in the history of CASP, the experimentalists not only highlighted that computational models can accurately reproduce the most critical structural features observed in their targets, but also envisaged that models could serve as a guidance for further studies of biologically‐relevant properties of proteins.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 20-12-2019
Abstract: The ligands bound by γδ T cell receptors (TCRs) are less well characterized than those of their αβ TCR cousins, which are antigens presented by major histocompatibility complex (MHC) and related proteins. Le Nours et al. identified a phenotypically erse γδ T cell subset in human tissues that reacts to MHC-related protein 1 (MR1), which presents vitamin B derivatives. A crystal structure of a γδ TCR–MR1–antigen complex revealed that some of these TCRs can bind underneath the MR1 antigen-binding cleft instead of recognizing the presented antigen. This work thus uncovers an additional ligand for γδ T cells and reconceptualizes the nature of T cell antigen recognition. Science , this issue p. 1522
Publisher: Elsevier BV
Date: 06-2019
DOI: 10.1016/J.STR.2019.03.009
Abstract: YjbH is a bacterial adaptor protein required for efficient proteolysis of the RNA polymerase-binding transcription factor Spx by the ClpXP protease. We report the structure of YjbH in complex with Spx. YjbH comprises a DsbA-like thioredoxin domain connected via a linker to a C-terminal domain reminiscent of the winged helix-turn-helix fold. The interaction between YjbH and Spx involves a large surface area. Binding to YjbH stabilizes the C-terminal ClpX recognition region of Spx. We show that mutation of critical YjbH contact residues abrogates Spx recognition. Small-angle X-ray scattering and hydrogen-deuterium exchange mass spectrometry analyses determined the existence of a stable heterodimeric complex in solution and provide evidence that binding of Spx to YjbH reduces the overall conformational flexibility of Spx. Our findings provide insights into the molecular basis for Spx recognition and suggest a model for how YjbH stabilizes Spx and displays the C terminus of Spx for engagement by ClpXP.
Publisher: Proceedings of the National Academy of Sciences
Date: 19-02-2013
Abstract: The origin and biological role of dynamic motions of folded enzymes is not yet fully understood. In this study, we examine the molecular determinants for the dynamic motions within the β-barrel of superoxide dismutase 1 (SOD1), which previously were implicated in allosteric regulation of protein maturation and also pathological misfolding in the neurodegenerative disease amyotrophic lateral sclerosis. Relaxation-dispersion NMR, hydrogen/deuterium exchange, and crystallographic data show that the dynamic motions are induced by the buried H43 side chain, which connects the backbones of the Cu ligand H120 and T39 by a hydrogen-bond linkage through the hydrophobic core. The functional role of this highly conserved H120–H43–T39 linkage is to strain H120 into the correct geometry for Cu binding. Upon elimination of the strain by mutation H43F, the apo protein relaxes through hydrogen-bond swapping into a more stable structure and the dynamic motions freeze out completely. At the same time, the holo protein becomes energetically penalized because the twisting back of H120 into Cu-bound geometry leads to burial of an unmatched backbone carbonyl group. The question then is whether this coupling between metal binding and global structural motions in the SOD1 molecule is an adverse side effect of evolving viable Cu coordination or plays a key role in allosteric regulation of biological function, or both?
Publisher: Proceedings of the National Academy of Sciences
Date: 29-11-2021
Abstract: Alongside αβ T cells and B cells, γδ T cells comprise a major component of the adaptive immune system, although a lack of bona fide ligands has hindered understanding of their function. γδ T cells are key mediators of epithelial immune surveillance and have a purported capacity for employing erse ligand engagement mechanisms beyond the dogmas of conventional αβ T cell–human leukocyte antigen restriction. Here, we found blood- and gut-resident Vδ1/2 − γδ T cells bound to MR1 tetramers in a metabolite-independent mechanism distinct from mucosal-associated invariant T cells and provide insight into a unique antibody-like MR1 recognition mode. This reshapes our understanding of the ligand recognition principles of γδ T cells and how they differ from αβ T cells.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 03-07-2020
DOI: 10.1126/SCIIMMUNOL.ABC9492
Abstract: A rare human allele renders MR1 unable to present microbially derived antigen, resulting in a selective loss of MAIT cells.
Location: United Kingdom of Great Britain and Northern Ireland
Location: United Kingdom of Great Britain and Northern Ireland
Start Date: 01-2022
End Date: 01-2025
Amount: $443,312.00
Funder: Australian Research Council
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