The Experts below are selected from a list of 327 Experts worldwide ranked by ideXlab platform
David Smil - One of the best experts on this subject based on the ideXlab platform.
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identification and characterization of the first fragment hits for setdb1 tudor domain
Bioorganic & Medicinal Chemistry, 2019Co-Authors: Philippe Mader, Rodrigo Mendozasanchez, A Dong, Elena Dobrovetsky, Victoria B Corless, Sean K Liew, S Houliston, Renato Ferreira De Freitas, A Iqbal, David SmilAbstract:Abstract SET domain bifurcated protein 1 (SETDB1) is a human histone-Lysine methyltransferase which is amplified in human cancers and was shown to be crucial in the growth of non-small and small cell lung carcinoma. In addition to its catalytic domain, SETDB1 harbors a unique tandem tudor domain which recognizes histone sequences containing both methylated and acetylated Lysines, and likely contributes to its localization on chromatin. Using X-ray crystallography and NMR spectroscopy fragment screening approaches, we have identified the first small molecule fragment hits that bind to histone peptide binding groove of the Tandem Tudor Domain (TTD) of SETDB1. Herein, we describe the binding modes of these fragments and analogues and the biophysical characterization of key compounds. These confirmed small molecule fragments will inform the development of potent antagonists of SETDB1 interaction with histones.
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identification and characterization of the first fragment hits for setdb1 tudor domain
bioRxiv, 2019Co-Authors: Philippe Mader, Rodrigo Mendozasanchez, A Dong, Elena Dobrovetsky, Victoria B Corless, Sean K Liew, S Houliston, Ferreira R De Freitas, A Iqbal, David SmilAbstract:ABSTRACT SET domain bifurcated protein 1 (SETDB1) is a human histone-Lysine methyltransferase, which is amplified in human cancers and was shown to be crucial in the growth of non-small and small cell lung carcinoma. In addition to its catalytic domain, SETDB1 harbors a unique tandem tudor domain which recognizes histone sequences containing both methylated and acetylated Lysines, and likely contributes to its localization on chromatin. Using X-ray crystallography and NMR spectroscopy fragment screening approaches, we have identified the first small molecule fragment hits that bind to histone peptide binding groove of the TTD of SETDB1. Herein, we describe the binding modes of these fragments and analogues and the biophysical characterization of key compounds. These confirmed small molecule fragments will inform the development of potent antagonists of SETDB1 interaction with histones.
Makoto Nishiyama - One of the best experts on this subject based on the ideXlab platform.
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crystal structure of lysk an enzyme catalyzing the last step of Lysine biosynthesis in thermus thermophilus in complex with Lysine insight into the mechanism for recognition of the amino group carrier protein lysw
Biochemical and Biophysical Research Communications, 2017Co-Authors: Satomi Fujita, Ayako Yoshida, Fumihito Hasebe, Takeo Tomita, Tomohisa Kuzuyama, Makoto NishiyamaAbstract:Abstract LysK is an M20 peptidase family enzyme that hydrolyzes the isopeptide bond between the carrier protein LysW and Lysine in order to release Lysine, which is the last step of Lysine biosynthesis in Thermus thermophilus . In the present study, we determined the crystal structure of LysK in complex with Lysine at a resolution of 2.4 A. The α-amino group of the bound Lysine was oriented toward the catalytic center, which was composed of the residues coordinating divalent metal ions for the hydrolysis of the isopeptide bond. An 11 A-long path was observed from the active site binding Lysine to the protein surface, which may be responsible for recognizing the C-terminal extension domain of LysW with the conserved EDWGE sequence. A positively-charged surface region was detected around the exit of the path, similar to other Lysine biosynthetic enzymes using LysW as the carrier protein. Mutational studies of the surface residues provided a plausible model for the electrostatic interaction with LysW.
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mechanism of substrate recognition and insight into feedback inhibition of homocitrate synthase from thermus thermophilus
Journal of Biological Chemistry, 2010Co-Authors: T Okada, Takeo Tomita, Tomohisa Kuzuyama, Asri Peni Wulandari, Makoto NishiyamaAbstract:Homocitrate synthase (HCS) catalyzes aldol-type condensation of acetyl coenzyme A (acetyl-CoA) and α-ketoglutarate (α-KG) to synthesize homocitrate (HC), which is the first and committed step in the Lysine biosynthetic pathway through α-aminoadipate. As known in most enzymes catalyzing the first reactions in amino acid biosynthetic pathways, HCS is regulated via feedback inhibition by the end product, Lysine. Here, we determined the crystal structures of HCS from Thermus thermophilus complexed with α-KG, HC, or Lysine. In the HC complex, the C1-carboxyl group of HC, which is derived from acetyl-CoA, is hydrogen-bonded with His-292* from another subunit (indicated by the asterisk), indicating direct involvement of this residue in the catalytic mechanism of HCS. The crystal structure of HCS complexed with Lysine showed that Lysine is bound to the active site with rearrangement of amino acid residues in the substrate-binding site, which accounts for the competitive inhibition by Lysine with α-KG. Comparison between the structures suggests that His-72, which is conserved in Lysine-sensitive HCSs and binds the C5-carboxyl group of α-KG, serves as a switch for the conformational change. Replacement of His-72 by leucine made HCS resistant to Lysine inhibition, demonstrating the regulatory role of this conserved residue.
Philippe Mader - One of the best experts on this subject based on the ideXlab platform.
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identification and characterization of the first fragment hits for setdb1 tudor domain
Bioorganic & Medicinal Chemistry, 2019Co-Authors: Philippe Mader, Rodrigo Mendozasanchez, A Dong, Elena Dobrovetsky, Victoria B Corless, Sean K Liew, S Houliston, Renato Ferreira De Freitas, A Iqbal, David SmilAbstract:Abstract SET domain bifurcated protein 1 (SETDB1) is a human histone-Lysine methyltransferase which is amplified in human cancers and was shown to be crucial in the growth of non-small and small cell lung carcinoma. In addition to its catalytic domain, SETDB1 harbors a unique tandem tudor domain which recognizes histone sequences containing both methylated and acetylated Lysines, and likely contributes to its localization on chromatin. Using X-ray crystallography and NMR spectroscopy fragment screening approaches, we have identified the first small molecule fragment hits that bind to histone peptide binding groove of the Tandem Tudor Domain (TTD) of SETDB1. Herein, we describe the binding modes of these fragments and analogues and the biophysical characterization of key compounds. These confirmed small molecule fragments will inform the development of potent antagonists of SETDB1 interaction with histones.
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identification and characterization of the first fragment hits for setdb1 tudor domain
bioRxiv, 2019Co-Authors: Philippe Mader, Rodrigo Mendozasanchez, A Dong, Elena Dobrovetsky, Victoria B Corless, Sean K Liew, S Houliston, Ferreira R De Freitas, A Iqbal, David SmilAbstract:ABSTRACT SET domain bifurcated protein 1 (SETDB1) is a human histone-Lysine methyltransferase, which is amplified in human cancers and was shown to be crucial in the growth of non-small and small cell lung carcinoma. In addition to its catalytic domain, SETDB1 harbors a unique tandem tudor domain which recognizes histone sequences containing both methylated and acetylated Lysines, and likely contributes to its localization on chromatin. Using X-ray crystallography and NMR spectroscopy fragment screening approaches, we have identified the first small molecule fragment hits that bind to histone peptide binding groove of the TTD of SETDB1. Herein, we describe the binding modes of these fragments and analogues and the biophysical characterization of key compounds. These confirmed small molecule fragments will inform the development of potent antagonists of SETDB1 interaction with histones.
Tobias C Walther - One of the best experts on this subject based on the ideXlab platform.
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native silac metabolic labeling of proteins in prototroph microorganisms based on Lysine synthesis regulation
Molecular & Cellular Proteomics, 2013Co-Authors: Florian Frohlich, Romain Christiano, Tobias C WaltherAbstract:Breakthroughs in proteomics (1–4) open up new possibilities for biological systems analysis. Central to these approaches is the necessity to accurately quantitate protein abundances. The most accurate quantitation is achieved by means of stable isotope labeling by amino acids in cell culture (SILAC)1 using heavy-isotope-containing amino acids (5). This approach has been widely used in many different experimental systems (6–11). SILAC relies on the metabolic incorporation of isotope-labeled Lysine and/or arginine into proteins. Samples from differently labeled cells are mixed and analyzed, for example, after one of them has been subjected to a different experimental condition. During sample preparation, proteins are digested to yield peptides containing one differentially labeled amino acid. As a consequence, mass spectrometry reveals “SILAC pairs” for each peptide, containing peaks corresponding to the unlabeled and the labeled peptides. The abundance ratio between the two reflects the different abundances of the protein in the starting samples. Lysine, which is commonly used in SILAC experiments, is an essential amino acid in higher eukaryotes that is obtained exclusively from food, but it can be synthesized in a tightly controlled fashion by plants, bacteria, and many fungi. Therefore, analysis of these prototroph organisms using SILAC has relied largely on mutants in Lysine biosynthesis. In contrast to other biochemical pathways of amino acid metabolism, Lysine biosynthesis occurs through distinct sets of reactions in different organisms that possess this biosynthetic capability. In bacteria, plants, and some fungi, Lysine is produced in nine enzymatically catalyzed steps via the diaminopimelate pathway from aspartate. In contrast, most fungi synthesize Lysine from the citrate cycle intermediate 2-oxoglutarate in ten steps, a pathway known as the α-aminoadipate pathway (12). Enzymes of this pathway are evolutionary conserved between such diverse species as S. pombe and S. cerevisiae, which are separated by 400 million years of evolution (13). Because of the specificity of Lysine biosynthesis pathways for bacteria and fungi, the enzymes participating in these reactions have become targets for the treatment of infections (14). An understanding of Lysine regulation is important for understanding the effects of such drugs, and it might also help in optimizing the industrial production of Lysine and other derived metabolites in these microorganisms. Both the diaminopimelate and the α-aminoadipate pathways of Lysine biosynthesis are exquisitely controlled: In S. cerevisiae, in which the regulation of Lysine metabolism has been studied in the most detail, the α-aminoadipate pathway is under combinatorial control. Product inhibition by Lysine controls homocitrate synthase, the first enzyme of the Lysine synthesis pathway, as well as the expression of key Lysine biosynthetic enzymes. Substrate feed-forward regulation by the intermediate l-2-aminoadipate-6-semialdehyde functions as a transcriptional co-activator, acting together with Lys14 to increase the amount of Lysine synthesis enzymes (15). Similarly, bacterial Lysine synthesis via the diaminopimelate pathway is regulated by a combination of transcriptional and post-transcriptional mechanisms. Specifically, dihydrodipicolinate synthase catalyzing the committing step of Lysine biosynthesis in bacteria is allosterically regulated by Lysine, and a number of genes in the pathways are transcriptionally regulated via Lysine repression (16). In order to determine the effects of Lysine in growth medium and to gain further insights into the biochemical regulation of Lysine synthesis pathways, we analyzed global proteome changes in different Lysine prototrophic model microorganisms in the presence or absence of Lysine. Our results reveal important differences in Lysine regulation between S. cerevisiae and S. pombe. Despite these differences, both organisms allowed us to exploit the regulation of Lysine biosynthesis for the development of a novel strategy for labeling prototrophic microorganisms. This strategy not only avoids complications of studying amino acid prototrophic mutant strains (e.g. when studying cellular metabolism), but also enables analysis of the large arsenal of mutants and systematic strain collections available for these model systems of cell biology, as well as industrially or medically important yeast and bacterial strains.
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proteome wide analysis of Lysine acetylation suggests its broad regulatory scope in saccharomyces cerevisiae
Molecular & Cellular Proteomics, 2012Co-Authors: Peter Henriksen, Brian T. Weinert, Sebastian A. Wagner, Satyan Sharma, Giedrė Bacinskaja, Michael Rehman, Andre H Juffer, Tobias C Walther, Michael Lisby, Chunaram ChoudharyAbstract:Post-translational modification of proteins by Lysine acetylation plays important regulatory roles in living cells. The budding yeast Saccharomyces cerevisiae is a widely used unicellular eukaryotic model organism in biomedical research. S. cerevisiae contains several evolutionary conserved Lysine acetyltransferases and deacetylases. However, only a few dozen acetylation sites in S. cerevisiae are known, presenting a major obstacle for further understanding the regulatory roles of acetylation in this organism. Here we use high resolution mass spectrometry to identify about 4000 Lysine acetylation sites in S. cerevisiae. Acetylated proteins are implicated in the regulation of diverse cytoplasmic and nuclear processes including chromatin organization, mitochondrial metabolism, and protein synthesis. Bioinformatic analysis of yeast acetylation sites shows that acetylated Lysines are significantly more conserved compared with nonacetylated Lysines. A large fraction of the conserved acetylation sites are present on proteins involved in cellular metabolism, protein synthesis, and protein folding. Furthermore, quantification of the Rpd3-regulated acetylation sites identified several previously known, as well as new putative substrates of this deacetylase. Rpd3 deficiency increased acetylation of the SAGA (Spt-Ada-Gcn5-Acetyltransferase) complex subunit Sgf73 on K33. This acetylation site is located within a critical regulatory domain in Sgf73 that interacts with Ubp8 and is involved in the activation of the Ubp8-containing histone H2B deubiquitylase complex. Our data provides the first global survey of acetylation in budding yeast, and suggests a wide-ranging regulatory scope of this modification. The provided dataset may serve as an important resource for the functional analysis of Lysine acetylation in eukaryotes.
Victoria B Corless - One of the best experts on this subject based on the ideXlab platform.
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identification and characterization of the first fragment hits for setdb1 tudor domain
Bioorganic & Medicinal Chemistry, 2019Co-Authors: Philippe Mader, Rodrigo Mendozasanchez, A Dong, Elena Dobrovetsky, Victoria B Corless, Sean K Liew, S Houliston, Renato Ferreira De Freitas, A Iqbal, David SmilAbstract:Abstract SET domain bifurcated protein 1 (SETDB1) is a human histone-Lysine methyltransferase which is amplified in human cancers and was shown to be crucial in the growth of non-small and small cell lung carcinoma. In addition to its catalytic domain, SETDB1 harbors a unique tandem tudor domain which recognizes histone sequences containing both methylated and acetylated Lysines, and likely contributes to its localization on chromatin. Using X-ray crystallography and NMR spectroscopy fragment screening approaches, we have identified the first small molecule fragment hits that bind to histone peptide binding groove of the Tandem Tudor Domain (TTD) of SETDB1. Herein, we describe the binding modes of these fragments and analogues and the biophysical characterization of key compounds. These confirmed small molecule fragments will inform the development of potent antagonists of SETDB1 interaction with histones.
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identification and characterization of the first fragment hits for setdb1 tudor domain
bioRxiv, 2019Co-Authors: Philippe Mader, Rodrigo Mendozasanchez, A Dong, Elena Dobrovetsky, Victoria B Corless, Sean K Liew, S Houliston, Ferreira R De Freitas, A Iqbal, David SmilAbstract:ABSTRACT SET domain bifurcated protein 1 (SETDB1) is a human histone-Lysine methyltransferase, which is amplified in human cancers and was shown to be crucial in the growth of non-small and small cell lung carcinoma. In addition to its catalytic domain, SETDB1 harbors a unique tandem tudor domain which recognizes histone sequences containing both methylated and acetylated Lysines, and likely contributes to its localization on chromatin. Using X-ray crystallography and NMR spectroscopy fragment screening approaches, we have identified the first small molecule fragment hits that bind to histone peptide binding groove of the TTD of SETDB1. Herein, we describe the binding modes of these fragments and analogues and the biophysical characterization of key compounds. These confirmed small molecule fragments will inform the development of potent antagonists of SETDB1 interaction with histones.
