Sally Boss

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Sub-single-turnover quantification of enzyme catalysis at ultrahigh throughput via a versatile NAD(P)H coupled assay in microdroplets

Enzyme engineering and discovery are crucial for a future sustainable bioeconomy, and harvesting ... more Enzyme engineering and discovery are crucial for a future sustainable bioeconomy, and harvesting new biocatalysts from large libraries through directed evolution or functional metagenomics requires accessible, rapid assays. Ultra-high throughput screening can often require an optical readout, leading to the use of model substrates that may not accurately report on activity for the target reaction and may require bespoke synthesis. In contrast, coupled assays represent a modular ‘plug-and-play’ system, where any pairing of enzyme/substrate may be investigated, if the reaction can produce a common intermediate which links the catalytic reaction to a detection cascade readout. Here we establish a detection cascade, producing a fluorescent readout in response to NAD(P)H via glutathione reductase and a subsequent thiol-mediated uncaging reaction, with a 30 nM detection limit. We demonstrate its utility for the glycosidaseAxyAgu115A (producing monosaccharides from a natural biofuel feedst...

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Controlled Ligand Exchange Between Ruthenium Organometallic Cofactor Precursors and a Naïve Protein Scaffold Generates Artificial Metalloenzymes Catalysing Transfer Hydrogenation

Angewandte Chemie, Mar 26, 2021

Many natural metalloenzymes assemble from proteins and biosynthesised complexes,generating potent... more Many natural metalloenzymes assemble from proteins and biosynthesised complexes,generating potent catalysts by changing metal coordination. Here we adopt the same strategy to generate artificial metalloenzymes (ArMs) using ligand exchange to unmask catalytic activity.Bysystematically testing Ru II (h 6-arene)(bipyridine) complexes designed to facilitate the displacement of functionalised bipyridines,wedevelop af ast and robust procedure for generating new enzymes via ligand exchange in aprotein that has not evolved to bind such ac omplex. The resulting metal cofactors form peptidic coordination bonds but also retain an on-biological ligand. Tandem mass spectrometry and 19 FNMR spectroscopyw ere used to characterise the organometallic cofactors and identify the protein-derived ligands.B yi ntroduction of ruthenium cofactors into a4-helical bundle,t ransfer hydrogenation catalysts were generated that displayed a3 5-fold rate increase when compared to the respective small molecule reaction in solution.

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Unlocking the Full Evolutionary Potential of Artificial Metalloenzymes Through Direct Metal-Protein Coordination : A review of recent advances for catalyst development

Johnson Matthey Technology Review, Oct 1, 2020

, before moving to the University of Cambridge to begin a PhD on the synthesis and reactivity of ... more

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Use of a fluorinated probe to quantitatively monitor amino acid binding preferences of ruthenium(<scp>ii</scp>) arene complexes

Dalton Transactions, 2019

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Understanding the Speciation of Ruthenium Arene Complexes as Precursors to Unnatural Enzyme Cofactors

Ruthenium arene complexes have been extensively explored as metallo-pharmaceuticals and as small ... more Ruthenium arene complexes have been extensively explored as metallo-pharmaceuticals and as small molecule catalysts. Exploring the overlap between these areas, this thesis describes a body of work aimed at quantitatively understanding the biological speciation and catalytic behaviour of ruthenium arene complexes when exposed to the many potential Lewis basic ligands provided by protein scaffolds. Combining nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (MS), X-ray crystallography and other biophysical characterisation techniques, the speciation of ruthenium arene bipyridine complexes with small molecule amino acids, short peptides and whole proteins has been monitored. A 19F NMR spectroscopic method was developed to quantitively trace the preferred amino acid binding partners of ruthenium complexes coordinated to fluorinated ligands. Liquid chromatography mass spectrometry (LC-MS) was used to explore the ligand exchange behaviour between ruthenium complexes and pro...

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CCDC 1867670: Experimental Crystal Structure Determination

CCDC 1867668: Experimental Crystal Structure Determination

CCDC 1867666: Experimental Crystal Structure Determination

CCDC 1867664: Experimental Crystal Structure Determination

Controlled Ligand Exchange Between Ruthenium Organometallic Cofactor Precursors and a Naïve Protein Scaffold Generates Artificial Metalloenzymes Catalysing Transfer Hydrogenation

Angewandte Chemie International Edition, 2021

Many natural metalloenzymes assemble from proteins and biosynthesised complexes, generating poten... more Many natural metalloenzymes assemble from proteins and biosynthesised complexes, generating potent catalysts by changing metal coordination. Here we adopt the same strategy to generate artificial metalloenzymes (ArMs) using ligand exchange to unmask catalytic activity. By systematically testing RuII(η6‐arene)(bipyridine) complexes designed to facilitate the displacement of functionalised bipyridines, we develop a fast and robust procedure for generating new enzymes via ligand exchange in a protein that has not evolved to bind such a complex. The resulting metal cofactors form peptidic coordination bonds but also retain a non‐biological ligand. Tandem mass spectrometry and 19F NMR spectroscopy were used to characterise the organometallic cofactors and identify the protein‐derived ligands. By introduction of ruthenium cofactors into a 4‐helical bundle, transfer hydrogenation catalysts were generated that displayed a 35‐fold rate increase when compared to the respective small molecule ...

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Use of a fluorinated probe to quantitatively monitor amino acid binding preferences of ruthenium(ii) arene complexes

Dalton Transactions, 2019

Speciation of Ru(ii) arene complexes in mixtures of amino acids with coordinating sidechains is e... more

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Tuning heavy metal compounds for anti-tumor activity: is diversity the key to ruthenium's success?

Future medicinal chemistry, 2009

This review aims to bring the reader up to date with the more recent ruthenium compounds that hav... more This review aims to bring the reader up to date with the more recent ruthenium compounds that have been synthesized and tested for their cytotoxicity. The chemistry of these transition metal complexes will be introduced and the basic principles that govern their common behavior outlined. The recent history of established compounds within this field will be presented alongside those that now represent the cutting-edge. The inherent variety within this class of compounds will lead the reader to appreciate their diversity and pose questions as to their similarities aside from the presence of a shared metal ion. This review aims to discuss and contextualize the state-of-the-art research within the context of the speculative advancement of this developing field. There is an evident need to specify the molecular and cellular targets of these drug molecules in order to ultimately elucidate their mode or modes of action. The evidence presented herein suggests that new avenues of research re...

Folates are potential ligands for ruthenium compounds in vivo

Dalton Trans., 2014

A labile ruthenium(ii) complex has been observed to chelate to folates under physiologically rele... more A labile ruthenium(ii) complex has been observed to chelate to folates under physiologically relevant conditions. The diastereomeric complexes formed would interfere with the one-carbon carrying role of folatein vivo. This highlights the importance of considering small molecules alongside macromolecules when determining the chemical origins of cytotoxicity of metallodrug candidates.

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Selective Lability of Ruthenium(II) Arene Amino Acid Complexes

Inorganic Chemistry, 2015

A series of organometallic complexes of the form [(PhH)Ru-(amino acid)] + have been synthesized u... more A series of organometallic complexes of the form [(PhH)Ru-(amino acid)] + have been synthesized using amino acids able to act as tridentate ligands. The straightforward syntheses gave enantiomerically pure complexes with two stereogenic centers due to the enantiopurity of the chelating ligands. Complexes were characterized in the solid-state and/or solution-state where the stability of the complex allowed. The propensity toward labilization of the coordinatively saturated complexes was investigated. The links between complex stability and structural features are very subtle. Nonetheless, H/D exchange rates of coordinated amino groups reveal more significant differences in reactivity linked to metallocycle ring size resulting in decreasing stability of the metallocycle as the amino acid side-chain length increases. The behavior of these systems in acid is unusual, apparently labilizing the carboxylate residue of the amino acid. This acid-catalyzed hemilability in an organometallic is relevant to the use of Ru(II) arenes in medicinal contexts due to the relatively low pH of cancerous cells.

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ChemInform Abstract: The Alkali and Coinage Metals

ChemInform, 2008

Oxygen scavenging by lithium zincates: the synthesis, structural characterisation and derivatisation of [Ph(2-C5H4N)N]2ZnRLi??nthf (R = But, Bun; n = 1, 2)

Journal of the Chemical Society, Dalton Transactions, 2002

... Sally R. Boss, Robert Haigh, David J. Linton and Andrew EH Wheatley*. Department of Chemistry... more

Toward an Understanding of the Oxygen Scavenging Properties of Lithium Zincates

Phosphorus, Sulfur, and Silicon and the Related Elements, 2004

The reaction of ZnMe2 with 2-pyridylamine [HN(2-C5H4N)Ph 1], LitBu and thereafter with dry air ha... more The reaction of ZnMe2 with 2-pyridylamine [HN(2-C5H4N)Ph 1], LitBu and thereafter with dry air has concomitantly yielded {[Ph(2-C5H4N)N]2ZnOMeLi·thf}2 (2) and [Ph(2-C5H4N)N]2Zn[(μ3-O) Bu]2(Li·thf)2 (3). The structure of 2 implies the insertion of oxygen into a [(R2N)2ZnMe]– ion. To probe this mechanism, we have prepared, characterized, and derivatized [Ph(2-C5H4N)N]2ZnRLi (R = nBu, n = 2, L = thf, 4a; R = tBu, n = 1, L = thf, 4b) (Figure 1). The sequential reaction of [Ph(2-C5H4N)N]2Zn with nBuLi, thf and O2 gives {[Ph(2C5H4N)N]2ZnOBuLi·nL}2 (n = 1, L = thf, 5), the structures of 4a and 5 strongly suggesting that oxygenation proceeds by insertion into the Zn C bond of an {[Ph(2-C5H4N)N]2ZnnBu}– ion. The treatment of [Ph(2-C5H4N)N]2Zn with tBuLi, thf, and O2 affords only the previously reported 3—this being in part rationalized in terms of the steric requirements of OtBu. Moreover, the structure of 3 is closely related to that of 5. Formally, this can be described in terms of the rearrangement of M O (M = Li, Zn) interactions in response to the presence or absence of a [Ph(2-C5H4N)N]2Zn moiety. Structural characterization of {[Ph(2-C5H4N)N]2ZnBuLi·dme}(6), {[Ph(2-C5H4N)N]2ZnOBuLi·0.5dme}2 (7), and [Ph(2-C5H4N)N]2 Zn[(μ3-O)Bu]2(Li2·dme) (8), further support this postulated oxygen insertion.

$Research paper thumbnail of Hydride Encapsulation by a Molecular Main-Group-Metal Cluster: Single-Crystal Neutron Diffraction Structure of [{Ph(2-C 5 H 4 N)N} 6 HLi 8 ] +$

Hydride Encapsulation by a Molecular Main-Group-Metal Cluster: Single-Crystal Neutron Diffraction Structure of [{Ph(2-C 5 H 4 N)N} 6 HLi 8 ] +

Organometallics, 2004

... Ser. 1978, No. 167, 93. (d) Albano, VG; Ciamician, G.; Ciani, G.; Martinengo, S.; Sironi, A. ... more

Encapsulation of hydride by molecular main group metal clusters: manipulating the source and coordination sphere of the interstitial ion

Dalton Trans., 2006

Synthesis of [{Me-4-C 6 H 4 (2-C 5 H 4 N)N} 6 HLi 8 ] + [(Bu t 2 AlMe 2) 2 Li]-•PhMe 5. Me 3 Al (... more

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New motifs in lithium zincate chemistry: a solid-state structural study of PhC(O)N(R)ZnR′2Li·2thf (R, R′ = alkyl, aryl) and [PhC(O)N(Ph)Li·thf]·[PhC(O)N(Ph)Zn(But)2Li·thf]

Dalton Transactions, 2003

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