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Ben Shen - One of the best experts on this subject based on the ideXlab platform.
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enediyne antitumor antibiotic maduropeptin biosynthesis featuring a c methyltransferase that acts on a coa tethered aromatic substrate
Journal of the American Chemical Society, 2010Co-Authors: Jianya Ling, Yinggang Luo, Shuangjun Lin, Geoffrey P Horsman, Shengxiong Huang, Ben ShenAbstract:The enediyne antitumor antibiotic maduropeptin (MDP) is produced by Actinomadura madurae ATCC 39144. The biosynthetic pathway for the 3,6-dimethylsalicylic acid moiety of the MDP chromophore is proposed to be comprised of four enzymes: MdpB, MdpB1, MdpB2, and MdpB3. Based on the previously characterized biosynthesis of the naphthoic acid moiety of Neocarzinostatin (NCS), we expected a biosynthetic pathway featuring carboxylic acid activation by the MdpB2 CoA ligase immediately before its coupling to an enediyne core intermediate. Surprisingly, the MDP aromatic acid biosynthetic pathway employs an unusual logic in which MdpB2-catalyzed CoA activation occurs before MdpB1-catalyzed C-methylation, demonstrating that MdpB1 is apparently unique in its ability to C-methylate a CoA-tethered aromatic acid. MdpB2 is a promiscuous CoA ligase capable of activating a variety of salicylic acid analogues, a property that could be potentially exploited to engineer MDP analogues.
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characterization of the epoxide hydrolase ncsf2 from the Neocarzinostatin biosynthetic gene cluster
Organic Letters, 2010Co-Authors: Shuangjun Lin, Geoffrey P Horsman, Ben ShenAbstract:Neocarzinostatin (1) biosynthesis is proposed to involve a vicinal diol intermediate. It is reported that NcsF2, one of two epoxide hydrolases encoded by the NCS gene cluster, catalyzes regiospecific addition of H2O to C-2 of both (R)- and (S)-styrene oxides to afford (R)- and (S)-1-phenyl-1,2-ethanediols, respectively, supporting its proposed role in 1 biosynthesis. (R)-1-Phenyl-1,2-ethanediol (87% yield and 99% ee) was obtained from (±)-styrene oxide hydrolysis by cocatalysis using NcsF2 and SgcF, the complementary epoxide hydrolase from the C-1027 biosynthetic pathway.
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molecular basis of substrate promiscuity for the sam dependent o methyltransferase ncsb1 involved in the biosynthesis of the enediyne antitumor antibiotic Neocarzinostatin
Biochemistry, 2009Co-Authors: Heather A. Cooke, Ben Shen, Yinggang Luo, Elizabeth L Guenther, Steven D. BrunerAbstract:The small molecule component of chromoprotein enediyne antitumor antibiotics is biosynthesized through a convergent route, incorporating amino acid, polyketide, and carbohydrate building blocks around a central enediyne hydrocarbon core. The naphthoic acid moiety of the enediyne Neocarzinostatin plays key roles in the biological activity of the natural product by interacting with both the carrier protein and duplex DNA at the site of action. We have previously described the in vitro characterization of an S-adenosylmethionine-dependent O-methyltransferase (NcsB1) in the Neocarzinostatin biosynthetic pathway [Luo, Y., Lin, S., Zhang, J., Cooke, H. A., Bruner, S. D., and Shen, B. (2008) J. Biol. Chem. 283, 14694−14702]. Here we provide a structural basis for NcsB1 activity, illustrating that the enzyme shares an overall architecture with a large family of S-adenosylmethionine-dependent proteins. In addition, NcsB1 represents the first enzyme to be structurally characterized in the biosynthetic pathway of ne...
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Regiospecific O-Methylation of Naphthoic Acids Catalyzed by NcsB1, an O-Methyltransferase Involved in the Biosynthesis of the Enediyne Antitumor Antibiotic Neocarzinostatin
The Journal of biological chemistry, 2008Co-Authors: Yinggang Luo, Heather A. Cooke, Jian Zhang, Steven D. Bruner, Shuangjun Lin, Ben ShenAbstract:Neocarzinostatin, a clinical anticancer drug, is the archetypal member of the chromoprotein family of enediyne antitumor antibiotics that are composed of a nonprotein chromophore and an apoprotein. The Neocarzinostatin chromophore consists of a nine-membered enediyne core, a deoxyaminosugar, and a naphthoic acid moiety. We have previously cloned and sequenced the Neocarzinostatin biosynthetic gene cluster and proposed that the biosynthesis of the naphthoic acid moiety and its incorporation into the Neocarzinostatin chromophore are catalyzed by five enzymes NcsB, NcsB1, NcsB2, NcsB3, and NcsB4. Here we report the biochemical characterization of NcsB1, unveiling that: (i) NcsB1 is an S-adenosyl-L-methionine-dependent O-methyltransferase; (ii) NcsB1 catalyzes regiospecific methylation at the 7-hydroxy group of its native substrate, 2,7-dihydroxy-5-methyl-1-naphthoic acid; (iii) NcsB1 also recognizes other dihydroxynaphthoic acids as substrates and catalyzes regiospecific O-methylation; and (iv) the carboxylate and its ortho-hydroxy groups of the substrate appear to be crucial for NcsB1 substrate recognition and binding, and O-methylation takes place only at the free hydroxy group of these dihydroxynaphthoic acids. These findings establish that NcsB1 catalyzes the third step in the biosynthesis of the naphthoic acid moiety of the Neocarzinostatin chromophore and further support the early proposal for the biosynthesis of the naphthoic acid and its incorporation into the Neocarzinostatin chromophore with free naphthoic acids serving as intermediates. NcsB1 represents another opportunity that can now be exploited to produce novel Neocarzinostatin analogs by engineering Neocarzinostatin biosynthesis or applying directed biosynthesis strategies.
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a phosphopantetheinylating polyketide synthase producing a linear polyene to initiate enediyne antitumor antibiotic biosynthesis
Proceedings of the National Academy of Sciences of the United States of America, 2008Co-Authors: Jian Zhang, Steven G. Van Lanen, Wen Liu, Pieter C Dorrestein, Neil L Kelleher, Ben ShenAbstract:The enediynes, unified by their unique molecular architecture and mode of action, represent some of the most potent anticancer drugs ever discovered. The biosynthesis of the enediyne core has been predicted to be initiated by a polyketide synthase (PKS) that is distinct from all known PKSs. Characterization of the enediyne PKS involved in C-1027 (SgcE) and Neocarzinostatin (NcsE) biosynthesis has now revealed that (i) the PKSs contain a central acyl carrier protein domain and C-terminal phosphopantetheinyl transferase domain; (ii) the PKSs are functional in heterologous hosts, and coexpression with an enediyne thioesterase gene produces the first isolable compound, 1,3,5,7,9,11,13-pentadecaheptaene, in enediyne core biosynthesis; and (iii) the findings for SgcE and NcsE are likely shared among all nine-membered enediynes, thereby supporting a common mechanism to initiate enediyne biosynthesis.
Andrew G. Myers - One of the best experts on this subject based on the ideXlab platform.
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development of an enantioselective synthetic route to Neocarzinostatin chromophore and its use for multiple radioisotopic incorporation
ChemInform, 2002Co-Authors: Andrew G. Myers, Marlys Hammond, Philip M. Harrington, Ralf Glatthar, Elaine Y Kuo, Jun Liang, Scott E Schaus, Jia-ning XiangAbstract:A convergent, enantioselective synthetic route to the natural product Neocarzinostatin chromophore (1) is described. Synthesis of the chromophore aglycon (2) was targeted initially. Chemistry previously developed for the synthesis of a Neocarzinostatin core model (4) failed in the requisite 1,3-transposition of an allylic silyl ether when applied toward the preparation of 2 with use of the more highly oxygenated substrates 27 and 54. An alternative synthetic plan was therefore developed, based upon a proposed reduction of the epoxy alcohol 58 to form the aglycon 2, a transformation that was achieved in a novel manner, using a combination of the reagents triphenylphosphine, iodine, and imidazole. The successful route to 1 and 2 began with the convergent coupling of the epoxydiyne 15, obtained in 9 steps (43% overall yield) from d-glyceraldehyde acetonide, and the cyclopentenone (+)-14, prepared in one step (75−85% yield) from the prostaglandin intermediate (+)-16, affording the alcohol 22 in 80% yield and ...
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gene transcription analysis of saccharomyces cerevisiae exposed to Neocarzinostatin protein chromophore complex reveals evidence of dna damage a potential mechanism of resistance and consequences of prolonged exposure
Proceedings of the National Academy of Sciences of the United States of America, 2001Co-Authors: Scott E Schaus, Duccio Cavalieri, Andrew G. MyersAbstract:The natural product Neocarzinostatin (NCS), a protein-small molecule complex, exhibits potent antiproliferative activity in mammalian cells but has little apparent effect on the growth of the unicellular eukaryotic organism, Saccharomyces cerevisiae. Here, we show by whole-genome transcription profiling experiments that incubation of S. cerevisiae with NCS leads to dramatic and wide-ranging modifications in the expression profile of yeast genes. Approximately 18% of yeast transcripts are altered by 2-fold or more within 4 h of treatment with NCS. Analysis of the observed transcription profile provides evidence that yeast rapidly and continuously overexpress multiple DNA-damage repair genes during NCS exposure. Perhaps to meet the energetic requirements of continuous DNA-damage repair, yeast cells enter respiration upon prolonged exposure to NCS, although grown in nutrient-rich medium. The NCS protein component is readily transported into S. cerevisiae, as demonstrated by fluorescence microscopy of yeast treated with fluorescently labeled NCS. Transcription profiling experiments with Neocarzinostatin protein alone implicate a specific resistance mechanism in yeast that targets the NCS protein component, one involving the nonclassical export pathway. These experiments provide a detailed picture of the effects of exposure to NCS upon yeast and the mechanisms they engage as a response to this protein-small molecule DNA-damaging agent.
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concise synthesis of the bicyclic core of the chromoprotein antibiotics kedarcidin and Neocarzinostatin by transannular reductive cyclization of a tetrayne precursor
Tetrahedron Letters, 1998Co-Authors: Andrew G. Myers, Steven D GoldbergAbstract:Abstract The oxygenated [7.3.0]-bicyclododecadienediyne 8 is synthesized in 9 steps from l -dimethyl tartrate. In the key transformation, transannular reductive cyclization of the potassium salt of the tetrayne alcohol 7 with NaAIH(OCH 2 CH 2 (NCH 3 ) 2 ) 3 affords the bicyclic product 8 in 50–54% yield. This sequence provides a rapid entry into the strained bicyclic core structures of kedarcidin and Neocarzinostatin chromophores ( 1 and 2 , respectively).
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a comparison of dna cleavage by Neocarzinostatin chromophore and its aglycon evaluating the role of the carbohydrate residue
Journal of the American Chemical Society, 1997Co-Authors: Andrew G. Myers, Michael E Kort, Marlys HammondAbstract:Through a comparative analysis of the reactivity and DNA cleaving activity of Neocarzinostatin (NCS) chromophore (1) and the corresponding aglycon (2), we show that the carbohydrate residue (aminoglycoside) of 1 both accelerates the rate and improves the efficiency of DNA cleavage versus the aglycon (2), but does not appear to be a major determinant of the base specificity of DNA cleavage by 1. This stands in contrast to earlier findings with another enediyne antibiotic, calicheamicin γ1, where the carbohydrate residue was found to be a major determinant of the sequence specificity of DNA cleavage, in addition to playing a functional role in the reductive activation step. Thiol addition experiments with NCS aglycon (2) provide further evidence that the carbohydrate amino group of 1 functions as an internal base in the first step leading to DNA cleavage, thiol activation. Evidence is also presented supporting the proposal that NCS aglycon (2) is bound tightly and reversibly by the Neocarzinostatin binding ...
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ENANTIOSELECTIVE SYNTHESIS OF Neocarzinostatin CHROMOPHORE AGLYCON
Journal of the American Chemical Society, 1996Co-Authors: Andrew G. Myers, Marlys Hammond, Yusheng Wu, Jia-ning Xiang, Philip M. HarringtonAbstract:Neocarzinostatin is the first of the “enediyne” antitumor agents to be characterized and is further distinguished as the first chromoprotein antibiotic.1 The DNA-cleaving properties of the Neocarzinostatin protein-chromophore complex have been shown to reside solely within the chromophore component (1) which, in isolation, is exceedingly unstable.2,3 The strain, structural complexity, and, most importantly, high reactivity of the chromophore core define an extraordinarily challenging synthetic target. In this work, we describe an enantioselective route that provides for the first time Neocarzinostatin chromophore aglycon (2), a substance which proves to be even less stable than 1 and which, almost certainly, could not be derived from the parent antibiotic.
Jian Zhang - One of the best experts on this subject based on the ideXlab platform.
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Regiospecific O-Methylation of Naphthoic Acids Catalyzed by NcsB1, an O-Methyltransferase Involved in the Biosynthesis of the Enediyne Antitumor Antibiotic Neocarzinostatin
The Journal of biological chemistry, 2008Co-Authors: Yinggang Luo, Heather A. Cooke, Jian Zhang, Steven D. Bruner, Shuangjun Lin, Ben ShenAbstract:Neocarzinostatin, a clinical anticancer drug, is the archetypal member of the chromoprotein family of enediyne antitumor antibiotics that are composed of a nonprotein chromophore and an apoprotein. The Neocarzinostatin chromophore consists of a nine-membered enediyne core, a deoxyaminosugar, and a naphthoic acid moiety. We have previously cloned and sequenced the Neocarzinostatin biosynthetic gene cluster and proposed that the biosynthesis of the naphthoic acid moiety and its incorporation into the Neocarzinostatin chromophore are catalyzed by five enzymes NcsB, NcsB1, NcsB2, NcsB3, and NcsB4. Here we report the biochemical characterization of NcsB1, unveiling that: (i) NcsB1 is an S-adenosyl-L-methionine-dependent O-methyltransferase; (ii) NcsB1 catalyzes regiospecific methylation at the 7-hydroxy group of its native substrate, 2,7-dihydroxy-5-methyl-1-naphthoic acid; (iii) NcsB1 also recognizes other dihydroxynaphthoic acids as substrates and catalyzes regiospecific O-methylation; and (iv) the carboxylate and its ortho-hydroxy groups of the substrate appear to be crucial for NcsB1 substrate recognition and binding, and O-methylation takes place only at the free hydroxy group of these dihydroxynaphthoic acids. These findings establish that NcsB1 catalyzes the third step in the biosynthesis of the naphthoic acid moiety of the Neocarzinostatin chromophore and further support the early proposal for the biosynthesis of the naphthoic acid and its incorporation into the Neocarzinostatin chromophore with free naphthoic acids serving as intermediates. NcsB1 represents another opportunity that can now be exploited to produce novel Neocarzinostatin analogs by engineering Neocarzinostatin biosynthesis or applying directed biosynthesis strategies.
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a phosphopantetheinylating polyketide synthase producing a linear polyene to initiate enediyne antitumor antibiotic biosynthesis
Proceedings of the National Academy of Sciences of the United States of America, 2008Co-Authors: Jian Zhang, Steven G. Van Lanen, Wen Liu, Pieter C Dorrestein, Neil L Kelleher, Ben ShenAbstract:The enediynes, unified by their unique molecular architecture and mode of action, represent some of the most potent anticancer drugs ever discovered. The biosynthesis of the enediyne core has been predicted to be initiated by a polyketide synthase (PKS) that is distinct from all known PKSs. Characterization of the enediyne PKS involved in C-1027 (SgcE) and Neocarzinostatin (NcsE) biosynthesis has now revealed that (i) the PKSs contain a central acyl carrier protein domain and C-terminal phosphopantetheinyl transferase domain; (ii) the PKSs are functional in heterologous hosts, and coexpression with an enediyne thioesterase gene produces the first isolable compound, 1,3,5,7,9,11,13-pentadecaheptaene, in enediyne core biosynthesis; and (iii) the findings for SgcE and NcsE are likely shared among all nine-membered enediynes, thereby supporting a common mechanism to initiate enediyne biosynthesis.
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characterization of ncsb2 as a promiscuous naphthoic acid coenzyme a ligase integral to the biosynthesis of the enediyne antitumor antibiotic Neocarzinostatin
Journal of the American Chemical Society, 2007Co-Authors: Heather A. Cooke, Jian Zhang, Meghan A. Griffin, Koichi Nonaka, Steven G. Van Lanen, Ben Shen, Steven D. BrunerAbstract:The enediyne antitumor antibiotic Neocarzinostatin (NCS) is produced by Streptomyces carzinostaticus ATCC15944. The biosynthetic pathway for the naphthoic acid moiety (boxed) of the NCS chromophore (1) is proposed to comprise five enzymes: NcsB, NcsB1, NcsB2, NcsB3, NcsB4. Both in vivo and in vitro experiments were used to support the proposed pathway. NcsB2 was characterized for the ability to catalyze the activation of 2-hydroxy-7-methoxy-5-methyl-1-naphthoic acid (3) via a naphthoyl-AMP ester (5) and subsequent formation of the corresponding coenzyme A ester. The finding that NcsB2 exhibits promiscuous substrate specificity toward a diverse set of naphthoic acid analogues with 2-, 5-, or 7-substitutions presents an outstanding opportunity to produce novel analogues of 1 by engineering NCS biosynthesis.
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the Neocarzinostatin biosynthetic gene cluster from streptomyces carzinostaticus atcc 15944 involving two iterative type i polyketide synthases
Chemistry & Biology, 2005Co-Authors: Wen Liu, Jian Zhang, Koichi Nonaka, Steven G. Van Lanen, Liping Nie, Steven D Christenson, Juyun Bae, Emmanuel Zazopoulos, Chris M Farnet, Catherine F YangAbstract:The biosynthetic gene cluster for the enediyne antitumor antibiotic Neocarzinostatin (NCS) was localized to 130 kb continuous DNA from Streptomyces carzinostaticus ATCC15944 and confirmed by gene inactivation. DNA sequence analysis of 92 kb of the cloned region revealed 68 open reading frames (ORFs), 47 of which were determined to constitute the NCS cluster. Sequence analysis of the genes within the NCS cluster suggested dNDP-D-mannose as a precursor for the deoxy aminosugar, revealed two distinct type I polyketide synthases (PKSs), and supported a convergent model for NCS chromophore biosynthesis from the deoxy aminosugar, naphthoic acid, and enediyne core building blocks. These findings shed light into deoxysugar biosynthesis, further support the iterative type I PKS paradigm for enediyne core biosynthesis, and unveil a mechanism for microbial polycyclic aromatic polyketide biosynthesis by an iterative type I PKS.
Steven D. Bruner - One of the best experts on this subject based on the ideXlab platform.
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molecular basis of substrate promiscuity for the sam dependent o methyltransferase ncsb1 involved in the biosynthesis of the enediyne antitumor antibiotic Neocarzinostatin
Biochemistry, 2009Co-Authors: Heather A. Cooke, Ben Shen, Yinggang Luo, Elizabeth L Guenther, Steven D. BrunerAbstract:The small molecule component of chromoprotein enediyne antitumor antibiotics is biosynthesized through a convergent route, incorporating amino acid, polyketide, and carbohydrate building blocks around a central enediyne hydrocarbon core. The naphthoic acid moiety of the enediyne Neocarzinostatin plays key roles in the biological activity of the natural product by interacting with both the carrier protein and duplex DNA at the site of action. We have previously described the in vitro characterization of an S-adenosylmethionine-dependent O-methyltransferase (NcsB1) in the Neocarzinostatin biosynthetic pathway [Luo, Y., Lin, S., Zhang, J., Cooke, H. A., Bruner, S. D., and Shen, B. (2008) J. Biol. Chem. 283, 14694−14702]. Here we provide a structural basis for NcsB1 activity, illustrating that the enzyme shares an overall architecture with a large family of S-adenosylmethionine-dependent proteins. In addition, NcsB1 represents the first enzyme to be structurally characterized in the biosynthetic pathway of ne...
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Regiospecific O-Methylation of Naphthoic Acids Catalyzed by NcsB1, an O-Methyltransferase Involved in the Biosynthesis of the Enediyne Antitumor Antibiotic Neocarzinostatin
The Journal of biological chemistry, 2008Co-Authors: Yinggang Luo, Heather A. Cooke, Jian Zhang, Steven D. Bruner, Shuangjun Lin, Ben ShenAbstract:Neocarzinostatin, a clinical anticancer drug, is the archetypal member of the chromoprotein family of enediyne antitumor antibiotics that are composed of a nonprotein chromophore and an apoprotein. The Neocarzinostatin chromophore consists of a nine-membered enediyne core, a deoxyaminosugar, and a naphthoic acid moiety. We have previously cloned and sequenced the Neocarzinostatin biosynthetic gene cluster and proposed that the biosynthesis of the naphthoic acid moiety and its incorporation into the Neocarzinostatin chromophore are catalyzed by five enzymes NcsB, NcsB1, NcsB2, NcsB3, and NcsB4. Here we report the biochemical characterization of NcsB1, unveiling that: (i) NcsB1 is an S-adenosyl-L-methionine-dependent O-methyltransferase; (ii) NcsB1 catalyzes regiospecific methylation at the 7-hydroxy group of its native substrate, 2,7-dihydroxy-5-methyl-1-naphthoic acid; (iii) NcsB1 also recognizes other dihydroxynaphthoic acids as substrates and catalyzes regiospecific O-methylation; and (iv) the carboxylate and its ortho-hydroxy groups of the substrate appear to be crucial for NcsB1 substrate recognition and binding, and O-methylation takes place only at the free hydroxy group of these dihydroxynaphthoic acids. These findings establish that NcsB1 catalyzes the third step in the biosynthesis of the naphthoic acid moiety of the Neocarzinostatin chromophore and further support the early proposal for the biosynthesis of the naphthoic acid and its incorporation into the Neocarzinostatin chromophore with free naphthoic acids serving as intermediates. NcsB1 represents another opportunity that can now be exploited to produce novel Neocarzinostatin analogs by engineering Neocarzinostatin biosynthesis or applying directed biosynthesis strategies.
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characterization of ncsb2 as a promiscuous naphthoic acid coenzyme a ligase integral to the biosynthesis of the enediyne antitumor antibiotic Neocarzinostatin
Journal of the American Chemical Society, 2007Co-Authors: Heather A. Cooke, Jian Zhang, Meghan A. Griffin, Koichi Nonaka, Steven G. Van Lanen, Ben Shen, Steven D. BrunerAbstract:The enediyne antitumor antibiotic Neocarzinostatin (NCS) is produced by Streptomyces carzinostaticus ATCC15944. The biosynthetic pathway for the naphthoic acid moiety (boxed) of the NCS chromophore (1) is proposed to comprise five enzymes: NcsB, NcsB1, NcsB2, NcsB3, NcsB4. Both in vivo and in vitro experiments were used to support the proposed pathway. NcsB2 was characterized for the ability to catalyze the activation of 2-hydroxy-7-methoxy-5-methyl-1-naphthoic acid (3) via a naphthoyl-AMP ester (5) and subsequent formation of the corresponding coenzyme A ester. The finding that NcsB2 exhibits promiscuous substrate specificity toward a diverse set of naphthoic acid analogues with 2-, 5-, or 7-substitutions presents an outstanding opportunity to produce novel analogues of 1 by engineering NCS biosynthesis.
Stephen Caddick - One of the best experts on this subject based on the ideXlab platform.
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Evaluating the use of Apo-Neocarzinostatin as a cell penetrating protein.
Protein Engineering Design and Selection, 2013Co-Authors: Paul Moody, Fabienne Burlina, Stephen R Martin, Rachel E Morgan, John Offer, Mark E B Smith, Justin E Molloy, Stephen CaddickAbstract:Protein-ligand complex Neocarzinostatin (NCS) is a small, thermostable protein-ligand complex that is able to deliver its ligand cargo into live mammalian cells where it induces DNA damage. Apo-NCS is able to functionally display complementarity determining regions loops, and has been hypothesised to act as a cell-penetrating protein, which would make it an ideal scaffold for cell targeting, and subsequent intracellular delivery of small-molecule drugs. In order to evaluate apo-NCS as a cell penetrating protein, we have evaluated the efficiency of its internalisation into live HeLa cells using matrix-assisted laser-desorption ionization-time-of-flight mass spectrometry and fluorescence microscopy. Following incubation of cells with apo-NCS, we observed no evidence of internalisation.
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An Efficient Synthesis of Epoxydiynes and a Key Fragment of Neocarzinostatin Chromophore
Organic letters, 2007Co-Authors: Baker, O. Thominet, H. Britton, Stephen CaddickAbstract:A key structural feature of the Neocarzinostatin chromophore is a reactive epoxydiyne. We present here a new method for the preparation of epoxydiynes by the addition of an allenyl zinc bromide to a propargylic ketone.
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Protein-small molecule interactions in Neocarzinostatin, the prototypical enediyne chromoprotein antibiotic
CHEMBIOCHEM, 2007Co-Authors: Stephen CaddickAbstract:The enediyne chromoproteins ore a class of potent antitumour antibiotics comprising a 1:1 complex of a protein and a noncovalently bound chromophore. The protein is required to protect and transport the highly labile chromophore, which acts as the cytotoxic component by reacting with DNA leading to strand cleavage. A derivative of the best-studied member of this class, Neocarzinostatin (NCS), is currently in use as a chemotherapeutic in Japan. The application of the chromoproteins as therapeutics along with their unique mode of action has prompted wide-spread interest in this area. Notable developments include the discovery of non-natural ligands for the apoproteins and the observotion that multiple binding modes are available for these ligands in the binding site. Mutation studies on the apoproteins have revealed much about their stability and variability, and the w application of an in vitro evolution method has conferred ne binding specificity for unrelated ligands. These investigations hold great promise for the application of the apoproteins for drug-delivery, transport and stabilisation systems.
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Synthetic strategies to epoxydiynes and a key synthon of the Neocarzinostatin chromophore
Organic & Biomolecular Chemistry, 2007Co-Authors: Stephen CaddickAbstract:We present herein our recent efforts towards the synthesis of epoxydiynes which represent an unusual structural feature of the Neocarzinostatin chromophore. A number of different routes to these epoxydiynes have been explored with varying success. Ultimately a concise and convergent approach was developed, which involved the addition of an allenyl zinc bromide to propargylic ketones/aldehydes followed by epoxide formation. This new protocol enabled us to synthesise a fully elaborated epoxydiyne which will find application for our studies towards the total synthesis of the NCS chromophore.
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synthetic ligands for apo Neocarzinostatin
Journal of the American Chemical Society, 2006Co-Authors: Stephen Caddick, Frederick W Muskett, Rhys G Stoneman, Derek N WoolfsonAbstract:Neocarzinostatin (NCS) is a 1:1 complex of an enediyne chromophore (NCSChrom), non-covalently bound to an 11 kDa protein (apoNCS). We are exploring apoNCS as a generic protein system for sequestering small molecules for therapeutic applications. Here, we disclose a new flavone ligand 1 for apoNCS and present a high-resolution NMR structure of this ligand bound to apoNCS. This is the first high-resolution structure of a completely non-cognate ligand bound to the apoNCS protein. This work provides unambiguous evidence that a completely new class of ligand can bind specifically to apoNCS. Furthermore, the mode of binding is different than that of the naphthoate-based ligands, and for such a simple hydrophobic compound, the new ligand surprisingly binds specifically. This work indicates that apo-Neocarzinostatin has multiple selective and distinct binding modes for small-molecule cargo.
