ORCID Profile
0000-0003-0069-9408
Current Organisation
The University of Auckland
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Publisher: International Union of Crystallography (IUCr)
Date: 29-08-2014
DOI: 10.1107/S2053230X14014745
Abstract: The rumen of dairy cattle can be thought of as a large, stable fermentation vat and as such it houses a large and erse community of microorganisms. The bacterium Butyrivibrio proteoclasticus is a representative of a significant component of this microbial community. It is a xylan-degrading organism whose genome encodes a large number of open reading frames annotated as fibre-degrading enzymes. This suite of enzymes is essential for the organism to utilize the plant material within the rumen as a fuel source, facilitating its survival in this competitive environment. Xsa43E, a GH43 enzyme from B. proteoclasticus , has been structurally and functionally characterized. Here, the structure of selenomethionine-derived Xsa43E determined to 1.3 Å resolution using single-wavelength anomalous diffraction is reported. Xsa43E possesses the characteristic five-bladed β-propeller domain seen in all GH43 enzymes. GH43 enzymes can have a range of functions, and the functional characterization of Xsa43E shows it to be an arabinofuranosidase capable of cleaving arabinose side chains from short segments of xylan. Full functional and structural characterization of xylan-degrading enzymes will aid in creating an enzyme cocktail that can be used to completely degrade plant material into simple sugars. These molecules have a range of applications as starting materials for many industrial processes, including renewable alternatives to fossil fuels.
Publisher: Wiley
Date: 07-01-2010
DOI: 10.1002/PROT.22662
Abstract: The release of polysaccharide from the plant cell wall is a key process to release the stored energy from plant biomass. Within the ruminant digestive system, a host of commensal microorganisms speed the breakdown of plant cell matter releasing fermentable sugars. The presence of phenolic compounds, most notably ferulic acid (FA), esterified within the cell wall is thought to pose a significant impediment to the degradation of the plant cell wall. The structure of a FA esterase from the ruminant bacterium Butyrivibrio proteoclasticus has been determined in two different space groups, in both the apo-form, and the ligand bound form with FA located in the active site. The structure reveals a new lid domain that has no structural homologues in the PDB. The flexibility of the lid domain is evident by the presence of three different conformations adopted by different molecules in the crystals. In the FA-bound structures, these conformations show sequential binding and closing of the lid domain over the substrate. Enzymatic activity assays demonstrate a broad activity against plant-derived hemicellulose, releasing at least four aromatic compounds including FA, coumaric acid, coumarin-3-carboxylic acid, and cinnamic acid. The rumen is a complex ecosystem that efficiently degrades plant biomass and the genome of B. proteoclasticus contains greater than 130 enzymes, which are potentially involved in this process of which Est1E is the first to be well characterized.
Publisher: American Chemical Society (ACS)
Date: 19-02-2020
Publisher: Wiley
Date: 25-02-2013
DOI: 10.1002/PROT.24254
Abstract: Butyrivibrio proteoclasticus is a significant component of the microbial population of the rumen of dairy cattle. It is a xylan-degrading organism whose genome encodes a large number of open reading frames annotated as fiber-degrading enzymes. We have determined the three-dimensional structure of Est2A, an acetyl xylan esterase from B. proteoclasticus, at 2.1 Å resolution, along with the structure of an inactive mutant (H351A) at 2.0 Å resolution. The structure reveals two domains-a C-terminal SGNH domain and an N-terminal jelly-roll domain typical of CE2 family structures. The structures are accompanied by experimentally determined enzymatic parameters against two model substrates, para-nitrophenyl acetate and para-nitrophenyl butyrate. The suite of fiber-degrading enzymes produced by B. proteoclasticus provides a rich source of new enzymes of potential use in industrial settings.
Publisher: International Union of Crystallography (IUCr)
Date: 02-2018
DOI: 10.1107/S1600576717017101
Abstract: A new optimized size exclusion chromatography small-angle X-ray scattering (SEC-SAXS) system for biomolecular SAXS at the Australian Synchrotron SAXS/WAXS beamline has been developed. The compact configuration reduces s le dilution to maximize sensitivity. Coflow s le presentation allows an 11-fold increase in flux on s le without capillary fouling, improving throughput and data quality, which are now primarily limited by the full flux available on the beamline. Multi-wavelength fibre optic UV analysis in close proximity to the X-ray beam allows for accurate concentration determination for s les with known UV extinction coefficients and thus estimation of the molecular weight of the scattering particle from the forward X-ray scattering intensity. Fast-flow low-volume SEC columns provide s le throughput competitive with batch concentration series measurements, albeit with a concomitant reduction of potential resolution relative to lower flow rates and larger SEC columns. The performance of the system is demonstrated using a set of model proteins, and its utility to solve various challenges is illustrated with a erse suite of protein s les. These developments increase the quality and rigor of SEC-SAXS analysis and open new avenues for biomolecular solution SEC-SAXS studies that have been challenged by low s le yields, temporal instability, radiation sensitivity and complex mixtures.
Publisher: Elsevier BV
Date: 07-2015
Publisher: International Union of Crystallography (IUCr)
Date: 13-10-2011
DOI: 10.1107/S0108768111037542
Abstract: Distinctive diffuse scattering in the form of diffuse rings around Bragg positions has been observed in the diffraction patterns of a crystal of the N -terminal fragment of the Gag protein from Feline Foamy Virus. It is shown that these are caused by geometric frustration as molecules try to pack on the triangular b – c mesh of the space group P 6 1 22. In order to explain the strong diffuse scattering it is necessary for the crystal to contain occupational disorder such that each unit cell contains one or other of two different molecular arrangements, A and B . The frustration arises because the nearest-neighbour packing prefers neighbouring cells to be AB or BA , which cannot be achieved on all three sides of a triangle simultaneously. To explain the observation that reciprocal sections hk 5 n , where n = integer, contain only Bragg peaks it is necessary that A and B are identical molecular arrangements differing only by a translation of 0.2 c . The implications of the disorder for solving the structure of the protein by conventional techniques as well as the possibility of using the diffuse scattering for this purpose are discussed.
Publisher: Springer Science and Business Media LLC
Date: 12-02-2021
DOI: 10.1038/S41467-021-21191-7
Abstract: The life cycle of Baculoviridae family insect viruses depends on the viral protein kinase, PK-1, to phosphorylate the regulatory protein, p6.9, to induce baculoviral genome release. Here, we report the crystal structure of Cydia pomenella granulovirus PK-1, which, owing to its likely ancestral origin among host cell AGC kinases, exhibits a eukaryotic protein kinase fold. PK-1 occurs as a rigid dimer, where an antiparallel arrangement of the αC helices at the dimer core stabilizes PK-1 in a closed, active conformation. Dimerization is facilitated by C-lobe:C-lobe and N-lobe:N-lobe interactions between protomers, including the domain-swapping of an N-terminal helix that crowns a contiguous β-sheet formed by the two N-lobes. PK-1 retains a dimeric conformation in solution, which is crucial for catalytic activity. Our studies raise the prospect that parallel, side-to-side dimeric arrangements that lock kinase domains in a catalytically-active conformation could function more broadly as a regulatory mechanism among eukaryotic protein kinases.
No related grants have been discovered for David Goldstone.