ORCID Profile
0000-0001-9212-0461
Current Organisations
The University of Auckland
,
Helmholtz-Zentrum Dresden-Rossendorf, Institute of Radiopharmaceutical Cancer Research, Research site Leipzig
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Publications
Radiation Damage and Racemic Protein Crystallography Reveal the Unique Structure of the GASA/Snakin Protein Superfamily
Publisher: Wiley
Date: 04-05-2016
Abstract: Proteins from the GASA/snakin superfamily are common in plant proteomes and have erse functions, including hormonal crosstalk, development, and defense. One 63-residue member of this family, snakin-1, an antimicrobial protein from potatoes, has previously been chemically synthesized in a fully active form. Herein the 1.5 Å structure of snakin-1, determined by a novel combination of racemic protein crystallization and radiation-damage-induced phasing (RIP), is reported. Racemic crystals of snakin-1 and quasi-racemic crystals incorporating an unnatural 4-iodophenylalanine residue were prepared from chemically synthesized d- and l-proteins. Breakage of the C-I bonds in the quasi-racemic crystals facilitated structure determination by RIP. The crystal structure reveals a unique protein fold with six disulfide crosslinks, presenting a distinct electrostatic surface that may target the protein to microbial cell surfaces.
MCR-1: a promising target for structure-based design of inhibitors to tackle polymyxin resistance
Publisher: Elsevier BV
Date: 2019
DOI: 10.1016/J.DRUDIS.2018.07.004
Abstract: The spread of a novel mobile colistin resistance gene (mcr1) has jeopardised the use of polymyxins, last-resort antibiotics that are used increasingly to treat infections caused by multidrug-resistant (MDR) Gram-negative pathogens. In early 2017, the WHO reported the global spread of mcr1 within a few years after its initial discovery in China. The protein encoded by mcr1 is a putative 60-kDa phosphoethanolamine (pEtN) transferase, MCR-1, and has been studied extensively since its discovery. Herein, we present a comprehensive review of MCR-1 covering its structure, function, and mechanism, to call for the rational drug design of molecular inhibitors of MCR-1 to use in colistin-based combination therapies.
Radiosynthesis of racemic and enantiomerically pure (−)-[18F]flubatine—A promising PET radiotracer for neuroimaging of α4β2 nicotinic acetylcholine receptors
Publisher: Elsevier BV
Date: 04-2013
DOI: 10.1016/J.APRADISO.2013.01.002
Abstract: (-)-[(18)F]flubatine is a promising agent for visualization by PET of cerebral α4β2 nicotinic acetylcholine receptors (nAChRs), which are implicated in psychiatric and neurodegenerative disorders. Here, we describe a substantially improved two-step radiosynthesis strategy for (-)-[(18)F]flubatine, based on the nucleophilic radiofluorination of an enantiomerically pure precursor followed by deprotection of the intermediate. An extensive leaving group rotecting group library of precursors was tested. Application of a trimethylammonium-iodide precursor with a Boc-protecting group provided the best results: labeling efficiencies of 80-95%, RCY of 60±5%, radiochemical purity of >98%, and a specific activity of >350GBq/μmol. The radiosynthesis is easily transferable to an automated synthesis module.
Discovery and characterisation of hydrazines as inhibitors of the immune suppressive enzyme, indoleamine 2,3-dioxygenase 1 (IDO1)
Publisher: Elsevier BV
Date: 12-2013
DOI: 10.1016/J.BMC.2013.10.037
Abstract: Screening of a fragment library identified 2-hydrazinobenzothiazole as a potent inhibitor of indoleamine 2,3-dioxygenase 1 (IDO1), an enzyme expressed by tumours that suppresses the immune system. Spectroscopic studies indicated that 2-hydrazinobenzothiazole interacted with the IDO1 haem and in silico docking predicted that the interaction was through hydrazine. Subsequent studies of hydrazine derivatives identified phenylhydrazine (IC50=0.25 ± 0.07 μM) to be 32-fold more potent than 2-hydrazinobenzothiazole (IC50=8.0 ± 2.3 μM) in inhibiting rhIDO1 and that it inhibited cellular IDO1 at concentrations that were noncytotoxic to cells. Here, phenylhydrazine is shown to inhibit IDO1 through binding to haem.
Synthesis and biological evaluation of both enantiomers of [18F]flubatine, promising radiotracers with fast kinetics for the imaging of α4β2-nicotinic acetylcholine receptors
Publisher: Elsevier BV
Date: 2014
DOI: 10.1016/J.BMC.2013.12.011
Abstract: Both enantiomers of the epibatidine analogue flubatine display high affinity towards the α4β2 nicotinic acetylcholine receptor (nAChR) in vitro, accompanied by negligible interactions with erse off-target proteins. Extended single dose toxicity studies in rodent indicated a NOEL (No Observed Effect Level) of 6.2μg/kg for (-)-flubatine and 1.55μg/kg for (+)-flubatine. We developed syntheses for both flubatine enantiomers and their corresponding precursors for radiolabeling. The newly synthesized trimethylammonium precursors allowed for highly efficient (18)F-radiolabelling in radiochemical yields >60% and specific activities >750GBq/μmol, thus making the radioligands practical for clinical investigation.
High-Resolution Crystal Structure of Plant Carboxylesterase AeCXE1, from Actinidia eriantha, and Its Complex with a High-Affinity Inhibitor Paraoxon,
Publisher: American Chemical Society (ACS)
Date: 26-01-2007
DOI: 10.1021/BI062046W
Abstract: Carboxylesterases (CXEs) are widely distributed in plants, where they have been implicated in roles that include plant defense, plant development, and secondary metabolism. We have cloned, overexpressed, purified, and crystallized a carboxylesterase from the kiwifruit species Actinidia eriantha (AeCXE1). The structure of AeCXE1 was determined by X-ray crystallography at 1.4 A resolution. The crystal structure revealed that AeCXE1 is a member of the alpha/beta-hydrolase fold superfamily, most closely related structurally to the hormone-sensitive lipase subgroup. The active site of the enzyme, located in an 11 A deep hydrophobic gorge, contains the conserved catalytic triad residues Ser169, Asp276, and His306. Kinetic analysis using artificial ester substrates showed that the enzyme can hydrolyze a range of carboxylester substrates with acyl groups ranging from C2 to C16, with a preference for butyryl moieties. This preference was supported by the discovery of a three-carbon acyl adduct bound to the active site Ser169 in the native structure. AeCXE1 was also found to be inhibited by organophosphates, with paraoxon (IC50 = 1.1 muM) a more potent inhibitor than dimethylchlorophosphate (DMCP IC50 = 9.2 muM). The structure of AeCXE1 with paraoxon bound was determined at 2.3 A resolution and revealed that the inhibitor binds covalently to the catalytic serine residue, with virtually no change in the structure of the enzyme. The structural information for AeCXE1 provides a basis for addressing the wider functional roles of carboxylesterases in plants.
Investigations of the key macrolactamisation step in the synthesis of cyclic tetrapeptide pseudoxylallemycin A
Publisher: Royal Society of Chemistry (RSC)
Date: 2019
DOI: 10.1039/C9OB00227H
Abstract: During total synthesis of pseudoxylallemycin A, an unstable intermediate was observed and appeared to be reactivated by coupling reagent by-products.
Harnessing ester bond chemistry for protein ligation.
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C6CC09899A
Abstract: The hydrolysis potential of ester bonds in covalently cross-linking proteins is captured in our novel protein ligation technology. This new type of "molecular superglue" based on the spontaneously-formed Thr-Gln ester bonds found in cell-surface adhesins, affords a unique mechanism to both rationally assemble and disassemble complex protein nanomaterials.
Validating TDP1 as an Inhibition Target for the Development of Chemosensitizers for Camptothecin-Based Chemotherapy Drugs
Publisher: Springer Science and Business Media LLC
Date: 23-06-2021
Cover Picture: Radiation Damage and Racemic Protein Crystallography Reveal the Unique Structure of the GASA/Snakin Protein Superfamily (Angew. Chem. Int. Ed. 28/2016)
Publisher: Wiley
Date: 12-05-2016
Preparation of truncated orf virus entry fusion complex proteins by chemical synthesis
Publisher: Wiley
Date: 20-03-2014
DOI: 10.1002/PSC.2627
Abstract: Members of the Chordopoxvirinae subfamily possess an unusual 11 protein entry-fusion complex (EFC) that is highly conserved and present in all species. The mode of action of this EFC is unknown, and the interactions of the constituent proteins are uncharacterised. Here, we present the chemical synthesis of membrane domain truncated linear constructs of two EFC proteins in orf virus, ORFV036 and 049. By using Boc solid phase peptide synthesis and native chemical ligation methods, these truncated proteins have been readily prepared in milligram quantities. These robust synthetic protocols allow ready access to these polypeptides to facilitate biological studies.
Total chemical synthesis of an Orf virus protein, ORFV002, an inhibitor of the master gene regulator NF‐κB
Publisher: Wiley
Date: 03-2014
DOI: 10.1002/BIP.22445
Abstract: ORFV002 is a novel orf viral protein (117 Aa) that inhibits nuclear events through the regulation of the transcriptional activity of NF‐κB, a master regulator of human gene expression (Diel et al., J Virol 2011, 85, 264–275). It is identified as the first nuclear inhibitor of NF‐κB produced by orf virus (ORFV) and no homologues in other genera of the Chordopoxvirinae subfamily have been reported to date (Diel et al., J Virol 2011, 85, 264–275). Our molecular structure predictions suggest that ORFV002 may mimic part of IκB, an inhibitor and natural human partner of NF‐κB. Recent advances in total chemical synthesis of proteins have provided solutions in overcoming challenges of current recombinant methods of protein isolation for structure elucidation. Aided by Boc solid phase peptide synthesis and native chemical ligation, ORFV002 was successfully synthesized in multimilligram amounts in good yield and high purity. © 2013 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 102: 137–144, 2014.
Chemical synthesis of γ‐secretase activating protein using pseudoglutamines as ligation sites
Publisher: Wiley
Date: 2015
DOI: 10.1002/BIP.22600
Abstract: The chemical synthesis of analogue of a novel γ-secretase activating protein, which may play a pivotal role in the formation of amyloid peptides, the precursor to Alzheimer's disease, is described. The linear polypeptide sequence, consisting of 121 amino acids was assembled from four unprotected peptide building blocks using a convergent ligation-based synthesis. A strategic mutation of three glutamine residues to cysteine enabled the ligations, and the cysteines were subsequently converted to pseudoglutamines, to mimic the native glutamine. The full length unfolded protein was obtained in milligram amounts and was demonstrated to be homogeneous by liquid chromatography and mass spectrometry.
Structures and kinetics of Thermotoga maritima MetY reveal new insights into the predominant sulfurylation enzyme of bacterial methionine biosynthesis
Publisher: Elsevier BV
Date: 2021
Bioinspired Total Synthesis and Stereochemical Revision of the Fungal Metabolite Pestalospirane B
Publisher: American Chemical Society (ACS)
Date: 16-06-2017
DOI: 10.1021/ACS.ORGLETT.7B01371
Abstract: The total synthesis of both enantiomers of pestalospirane B, 2, has been achieved using a bioinspired tandem dimerization-spiroketalization reaction. Electronic circular dichroism (ECD) and X-ray analysis were used to revise the absolute stereochemistry of the natural product pestalospirane B from 3S, 3'S, 12R, 12'R to its enantiomer 3R, 3'R, 12S, 12'S.
Related Organisations
Helmholtz-Zentrum Dresden-Rossendorf, Institute Of Radiopharmaceutical Cancer Research, Research Site Leipzig
Location: Germany
Related Funding Activities
No related grants have been discovered for Christopher Squire.