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Kirsty J. McLean

Bio: Kirsty J. McLean is an academic researcher from University of Manchester. The author has contributed to research in topics: Mycobacterium tuberculosis & Gamma-ray burst. The author has an hindex of 37, co-authored 120 publications receiving 4736 citations. Previous affiliations of Kirsty J. McLean include Los Alamos National Laboratory & Elettra Sincrotrone Trieste.


Papers
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Journal ArticleDOI
TL;DR: The fundamental properties of P 450 BM3 are discussed and how progress with this model P450 has affected the authors' comprehension of P450 systems in general is discussed.

403 citations

Journal ArticleDOI
TL;DR: The catalytic potential of the P450s in organic biotransformations is the subject of this review—with emphasis on the breadth of P450 redox systems now recognised and the catalytic versatility of these biotechnologically important enzymes.

260 citations

Journal ArticleDOI
12 May 2005-Nature
TL;DR: In this paper, the authors reported a new optical component to internal shocks driven into the burst ejecta by variations of the inner engine, which is correlated with the prompt γ-rays, indicating a common origin for the optical light and the γrays.
Abstract: The prompt optical emission that arrives with the γ-rays from a cosmic γ-ray burst (GRB) is a signature of the engine powering the burst, the properties of the ultra-relativistic ejecta of the explosion, and the ejecta's interactions with the surroundings1,2,3,4,5. Until now, only GRB 990123 had been detected6 at optical wavelengths during the burst phase. Its prompt optical emission was variable and uncorrelated with the prompt γ-ray emission, suggesting that the optical emission was generated by a reverse shock arising from the ejecta's collision with surrounding material. Here we report prompt optical emission from GRB 041219a. It is variable and correlated with the prompt γ-rays, indicating a common origin for the optical light and the γ-rays. Within the context of the standard fireball model of GRBs, we attribute this new optical component to internal shocks driven into the burst ejecta by variations of the inner engine. The correlated optical emission is a direct probe of the jet isolated from the medium. The timing of the uncorrelated optical emission is strongly dependent on the nature of the medium.

203 citations

Journal ArticleDOI
TL;DR: The current state of knowledge of the diversity of these P450-redox partner fusion proteins is examined and their structural composition and evolutionary origins are explored.

198 citations

Journal ArticleDOI
TL;DR: When present at low concentrations in growth medium, azole antifungal drugs were shown to be potent inhibitors of the growth of Mycobacterium smegmatis and of Streptomyces strains, indicating that one or more of the P450s in these bacteria were viable drug targets.
Abstract: The genome sequence of Mycobacterium tuberculosis has revealed the presence of 20 different cytochrome P450 mono-oxygenases (P450s) within this organism, and subsequent genome sequences of other mycobacteria and of Streptomyces coelicolor have indicated that these actinomycetes also have large complements of P450s, pointing to important physiological roles for these enzymes. The actinomycete P450s include homologues of 14alpha-sterol demethylases, the targets for the azole class of drugs in yeast and fungi. Previously, this type of P450 was considered to be absent from bacteria. When present at low concentrations in growth medium, azole antifungal drugs were shown to be potent inhibitors of the growth of Mycobacterium smegmatis and of Streptomyces strains, indicating that one or more of the P450s in these bacteria were viable drug targets. The drugs econazole and clotrimazole were most effective against M. smegmatis (MIC values of <0.2 and 0.3 micro M, respectively) and were superior inhibitors of mycobacterial growth compared to rifampicin and isoniazid (which had MIC values of 1.2 and 36.5 micro M, respectively). In contrast to their effects on the actinomycetes, the azoles showed minimal effects on the growth of Escherichia coli, which is devoid of P450s. Azole drugs coordinated tightly to the haem iron in M. tuberculosis H37Rv P450s encoded by genes Rv0764c (the sterol demethylase CYP51) and Rv2276 (CYP121). However, the azoles had a higher affinity for M. tuberculosis CYP121, with K(d) values broadly in line with the MIC values for M. smegmatis. This suggested that CYP121 may be a more realistic target enzyme for the azole drugs than CYP51, particularly in light of the fact that an S. coelicolor DeltaCYP51 strain was viable and showed little difference in its sensitivity to azole drugs compared to the wild-type. If the azole drugs prove to inhibit a number of important P450s in M. smegmatis and S. coelicolor, then the likelihood of drug resistance developing in these species should be minimal. This suggests that azole drug therapy may provide a novel antibiotic strategy against strains of M. tuberculosis that have already developed resistance to isoniazid and other front-line drugs.

172 citations


Cited by
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Journal ArticleDOI
TL;DR: In this article, the interplay between these observations and theoretical models of the prompt gamma-ray burst and its afterglow is reviewed, and a model of the burst's origin and mechanism is proposed.
Abstract: Gamma-ray bursts are the most luminous explosions in the Universe, and their origin and mechanism are the focus of intense research and debate. More than three decades after their discovery, and after pioneering breakthroughs from space and ground experiments, their study is entering a new phase with the recently launched Swift satellite. The interplay between these observations and theoretical models of the prompt gamma-ray burst and its afterglow is reviewed.

1,294 citations

Journal Article
TL;DR: The highly automated PHENIX AutoBuild wizard is described, which can be applied equally well to phases derived from isomorphous/anomalous and molecular-replacement methods.
Abstract: Iterative model-building, structure refinement, and density modification with the PHENIX AutoBuild Wizard Thomas C. Terwilliger a* , Ralf W. Grosse-Kunstleve b , Pavel V. Afonine b , Nigel W. Moriarty b , Peter Zwart b , Li-Wei Hung a , Randy J. Read c , Paul D. Adams b* a b Los Alamos National Laboratory, Mailstop M888, Los Alamos, NM 87545, USA Lawrence Berkeley National Laboratory, One Cyclotron Road, Bldg 64R0121, Berkeley, CA 94720, USA. c Department of Haematology, University of Cambridge, Cambridge CB2 0XY, UK. * Email: terwill@lanl.gov or PDAdams@lbl.gov Running title: The PHENIX AutoBuild Wizard Abstract The PHENIX AutoBuild Wizard is a highly automated tool for iterative model- building, structure refinement and density modification using RESOLVE or TEXTAL model- building, RESOLVE statistical density modification, and phenix.refine structure refinement. Recent advances in the AutoBuild Wizard and phenix.refine include automated detection and application of NCS from models as they are built, extensive model completion algorithms, and automated solvent molecule picking. Model completion algorithms in the AutoBuild Wizard include loop-building, crossovers between chains in different models of a structure, and side-chain optimization. The AutoBuild Wizard has been applied to a set of 48 structures at resolutions ranging from 1.1 A to 3.2 A, resulting in a mean R-factor of 0.24 and a mean free R factor of 0.29. The R-factor of the final model is dependent on the quality of the starting electron density, and relatively independent of resolution. Keywords: Model building; model completion; macromolecular models; Protein Data Bank; structure refinement; PHENIX Introduction Iterative model-building and refinement is a powerful approach to obtaining a complete and accurate macromolecular model. The approach consists of cycles of building an atomic model based on an electron density map for a macromolecular structure, refining the structure, using the refined structure as a basis for improving the map, and building a new model. This type of approach has been carried out in a semi-automated fashion for many years, with manual model-building iterating with automated refinement (Jensen, 1997). More recently, with the development first of ARP/wARP (Perrakis et al., 1999), and later other procedures including RESOLVE iterative model-building and refinement (Terwilliger,

1,161 citations

Journal ArticleDOI
TL;DR: This work uses simulations with model lattice proteins to demonstrate how extra stability increases evolvability by allowing a protein to accept a wider range of beneficial mutations while still folding to its native structure.
Abstract: The biophysical properties that enable proteins to so readily evolve to perform diverse biochemical tasks are largely unknown. Here, we show that a protein’s capacity to evolve is enhanced by the mutational robustness conferred by extra stability. We use simulations with model lattice proteins to demonstrate how extra stability increases evolvability by allowing a protein to accept a wider range of beneficial mutations while still folding to its native structure. We confirm this view experimentally by mutating marginally stable and thermostable variants of cytochrome P450 BM3. Mutants of the stabilized parent were more likely to exhibit new or improved functions. Only the stabilized P450 parent could tolerate the highly destabilizing mutations needed to confer novel activities such as hydroxylating the antiinflammatory drug naproxen. Our work establishes a crucial link between protein stability and evolution. We show that we can exploit this link to discover protein functions, and we suggest how natural evolution might do the same.

1,085 citations

Journal ArticleDOI
TL;DR: In this article, the authors systematically analyze the possible physical processes that shape the properties of the early X-ray afterglow light curves and use the data to constrain various models.
Abstract: With the successful launch of the Swift Gamma-Ray Burst Explorer, a rich trove of early X-ray afterglow data has been collected by its onboard X-Ray Telescope (XRT). Some interesting features are emerging, including a distinct rapidly decaying component preceding the conventional afterglow component in many sources, a shallow decay component before the more "normal'' decay component observed in a good fraction of GRBs, and X-ray flares in nearly half of the afterglows. In this paper we systematically analyze the possible physical processes that shape the properties of the early X-ray afterglow light curves and use the data to constrain various models. We suggest that the steep decay component is consistent with the tail emission of the prompt gamma-ray bursts and/or the X-ray flares. This provides strong evidence that the prompt emission and afterglow emission are likely two distinct components, supporting the internal origin of the GRB prompt emission. The shallow decay segment observed in a group of GRBs suggests that very likely the forward shock keeps being refreshed for some time. This might be caused by either a long-lived central engine, or a wide distribution of the shell Lorentz factors, or else possibly the deceleration of a Poynting flux-dominated flow. X-ray flares suggest that the GRB central engine is very likely still active after the prompt gamma-ray emission is over, but with a reduced activity at later times. In some cases, the central engine activity even extends to days after the burst triggers. Analyses of early X-ray afterglow data reveal that GRBs are indeed highly relativistic events and that early afterglow data of many bursts, starting from the beginning of the XRT observations, are consistent with the afterglow emission from an ISM environment.

998 citations

Journal ArticleDOI
TL;DR: Although these proteins have properties that make them particularly attractive for engineering purposes, the large reservoir of P450 enzymes that collectively catalyze an astounding diversity of reactions suggests that P450 catalysis will develop into a highly useful technology.
Abstract: In chemical terms, the regio- and stereoselective hydroxylation of hydrocarbon C-H bonds is a very difficult transformation. Nevertheless, these reactions are deftly catalyzed by a variety of metalloenzymes, among which the most diverse are the many members of the cytochrome P450 family. Cytochrome P450 enzymes are found in most classes of organisms, including bacteria, fungi, plants, insects, and mammals. Thousands of such proteins are now known (http://drnelson.utmem.edu/cytochromeP450.html), including 57 in the human genome (1), 20 in Mycobacterium tuberculosis (2), 272 in Arabidopsis (3), and the amazing number of 457 in rice (4). The nomenclature for these enzymes is based on their sequence similarity when appropriately aligned, a somewhat arbitrary similarity cutoff (approximately >40% identity) being used to define members of a family and a higher cutoff (approximately >55% identity) members of a subfamily (5). Thus CYP3A4 corresponds to the fourth enzyme in family 3, subfamily A. This nomenclature allows the naming of enzymes without regard to their origin or specific properties. The mammalian, plant, and fungal proteins are commonly membrane bound and are relatively difficult to manipulate, but the bacterial proteins are usually soluble, monomeric proteins. For that reason, much of the early research on mechanisms of cytochrome P450 enzymes was carried out with bacterial enzymes, particularly with the prototypical enzyme CYP101 (P450cam) from Pseudomonas putida (6, 7). From a chemist's point of view, there is a particular interest in the thermophilic enzymes, which currently include CYP119 (8-10), P450st (11), CYP174A1 (12), and CYP231A2 (13). The thermal stability of these enzymes makes them attractive starting points for the development of industrially useful catalysts. In this context, particular attention has also focused on CYP102 (P450BM3), a self-sufficient enzyme from Bacillus megaterium in which the flavoprotein protein required for transfer of electrons from NADPH is fused to the hemoprotein (14). The resulting simplicity and high catalytic rate have led to extensive efforts to engineer this protein for practical catalytic purposes (15-19). Although these proteins have properties that make them particularly attractive for engineering purposes, the large reservoir of P450 enzymes that collectively catalyze an astounding diversity of reactions suggests that P450 catalysis will develop into a highly useful technology. The cytochrome P450 enzymes are defined by the presence in the proteins of a heme (iron protoporphyrin IX) prosthetic group coordinated on the proximal side by a thiolate ion (20, 21). This feature gives rise to the spectroscopic signature that defines these enzymes, as the thiolate-ligated ferrous-CO complex is characterized by a Soret absorption maximum at ∼450 nm (21). A thiolate-coordinated heme group is present in all P450 enzymes, although not all proteins with such coordination are members of this superfamily. One obvious exception, for example, is chloroperoxidase, which has a thiolate-coordinated heme group but normally catalyzes a very different reaction than the P450 enzymes (21-23). Although there are unusual P450 enzymes, such as the thromboxane and prostacyclin synthases (24), or CYP152 from Sphingomonas paucimobilis or Bacillus subtilis (25, 26), that normally utilize peroxides as substrates, the defining reaction for P450 enzymes is the reductive activation of molecular oxygen. In this reaction, one of the oxygen atoms of molecular oxygen is inserted into the substrate and the other oxygen atom is reduced to a molecule of water. With one exception to date (27, 28), the electrons required for this reduction of molecular oxygen derive from reduced pyridine nucleotides (NADH or NADPH). The overall equation for the reaction thus adheres to the formula: RH + NAD(P)H + O2 + H+ -> ROH + NAD(P)+ + H2O, where RH stands for a substrate with a hydroxylatable site. P450 enzymes therefore belong to the monooxygenase class of enzymes that only insert one of the oxygen atoms of molecular oxygen into their substrates. However, under appropriate circumstances or with specific substrates, other P450-catalyzed reactions can be observed, including desaturation, carbon-carbon bond scission, and carbon-carbon bond formation (29, 30). This review specifically focuses on P450-catalyzed hydrocarbon hydroxylation, the reaction that is most characteristic of P450 enzymes and that has been most extensively investigated. However, the principles that apply in these reactions also apply to other hydroxylation reactions, including those that occur on carbons adjacent to nitrogen, sulfur, or oxygen.

880 citations