The Experts below are selected from a list of 281250 Experts worldwide ranked by ideXlab platform
Matthew Cotten - One of the best experts on this subject based on the ideXlab platform.
-
increased resolution of african swine fever virus genome patterns based on profile hmms of Protein Domains
Virus Evolution, 2020Co-Authors: Charles Masembe, My V T Phan, David Robertson, Matthew CottenAbstract:African swine fever virus (ASFV), belonging to the Asfarviridae family, was originally described in Africa almost 100 years ago and is now spreading uncontrolled across Europe and Asia and threatening to destroy the domestic pork industry. Neither effective antiviral drugs nor protective vaccines are currently available. Efforts to understand the basis for viral pathogenicity and the development of attenuated potential vaccine strains are complicated by the large and complex nature of the ASFV genome. We report here a novel alignment-free method of documenting viral diversity based on profile hidden Markov model Domains on a genome scale. The method can be used to infer genomic relationships independent of genome alignments and also reveal ASFV genome sequence differences that determine the presence and characteristics of functional Protein Domains in the virus. We show that the method can quickly identify differences and shared patterns between virulent and attenuated ASFV strains and will be a useful tool for developing much-needed vaccines and antiviral agents to help control this virus. The tool is rapid to run and easy to implement, readily available as a simple Docker image.
-
increased resolution of african swine fever virus genome patterns based on profile hmm Protein Domains
bioRxiv, 2020Co-Authors: Charles Masembe, My V T Phan, David Robertson, Matthew CottenAbstract:African Swine Fever Virus (ASFV) was originally described in Africa almost 100 years ago and is now spreading uncontrolled across Europe and Asia and threatening to destroy the domestic pork industry. Neither effective antiviral drugs nor a protective vaccine are currently available. Efforts to understand the basis for viral pathogenicity and the development of attenuated potential vaccine strains are complicated by the large and complex ASFV genome. We report here a novel method of documenting viral diversity based on profile Hidden Markov Model Domains on a genome scale. The method can be used to infer genomic relationships independent of genome alignments and also reveal ASFV genome sequence differences that alter the presence of functional Protein Domains in the virus. We show that the method can quickly identify differences and shared patterns between virulent and attenuated ASFV strains and will be a useful tool for developing much-needed vaccines and antiviral agents to help control this virus. The tool is rapid to run and easy to implement, readily available as a simple Docker image.
Stephen P Jackson - One of the best experts on this subject based on the ideXlab platform.
-
detection of functional Protein Domains by unbiased genome wide forward genetic screening
Scientific Reports, 2018Co-Authors: Mareike Herzog, Fabio Puddu, Julia Coates, Nicola J Geisler, Josep V Forment, Stephen P JacksonAbstract:Establishing genetic and chemo-genetic interactions has played key roles in elucidating mechanisms by which certain chemicals perturb cellular functions. In contrast to gene disruption/depletion strategies to identify mechanisms of drug resistance, searching for point-mutational genetic suppressors that can identify separation- or gain-of-function mutations has been limited. Here, by demonstrating its utility in identifying chemical-genetic suppressors of sensitivity to the DNA topoisomerase I poison camptothecin or the poly(ADP-ribose) polymerase inhibitor olaparib, we detail an approach allowing systematic, large-scale detection of spontaneous or chemically-induced suppressor mutations in yeast or haploid mammalian cells in a short timeframe, and with potential applications in other haploid systems. In addition to applications in molecular biology research, this protocol can be used to identify drug targets and predict drug-resistance mechanisms. Mapping suppressor mutations on the primary or tertiary structures of Protein suppressor hits provides insights into functionally relevant Protein Domains. Importantly, we show that olaparib resistance is linked to missense mutations in the DNA binding regions of PARP1, but not in its catalytic domain. This provides experimental support to the concept of PARP1 trapping on DNA as the prime source of toxicity to PARP inhibitors, and points to a novel olaparib resistance mechanism with potential therapeutic implications.
-
detection of functional Protein Domains by unbiased genome wide forward genetic screening
bioRxiv, 2017Co-Authors: Mareike Herzog, Fabio Puddu, Julia Coates, Nicola J Geisler, Josep V Forment, Stephen P JacksonAbstract:Genetic and chemo-genetic interactions have played key roles in elucidating the molecular mechanisms by which certain chemicals perturb cellular functions. Many studies have employed gene knockout collections or gene disruption/depletion strategies to identify routes for evolving resistance to chemical agents. By contrast, searching for point-mutational genetic suppressors that can identify separation- or gain-of-function mutations, has been limited even in simpler, genetically amenable organisms such as yeast, and has not until recently been possible in mammalian cell culture systems. Here, by demonstrating its utility in identifying suppressors of cellular sensitivity to the drugs camptothecin or olaparib, we describe an approach allowing systematic, large-scale detection of spontaneous or chemically-induced suppressor mutations in yeast and in haploid mouse embryonic stem cells in a short timeframe, and with potential applications in essentially any other haploid system. In addition to its utility for molecular biology research, this protocol can be used to identify drug targets and to predict mechanisms leading to drug resistance. Mapping suppressor mutations on the primary sequence or three-dimensional structures of Protein suppressor hits provides insights into functionally relevant Protein Domains, advancing our molecular understanding of Protein functions, and potentially helping to improve drug design and applicability.
Josep V Forment - One of the best experts on this subject based on the ideXlab platform.
-
detection of functional Protein Domains by unbiased genome wide forward genetic screening
Scientific Reports, 2018Co-Authors: Mareike Herzog, Fabio Puddu, Julia Coates, Nicola J Geisler, Josep V Forment, Stephen P JacksonAbstract:Establishing genetic and chemo-genetic interactions has played key roles in elucidating mechanisms by which certain chemicals perturb cellular functions. In contrast to gene disruption/depletion strategies to identify mechanisms of drug resistance, searching for point-mutational genetic suppressors that can identify separation- or gain-of-function mutations has been limited. Here, by demonstrating its utility in identifying chemical-genetic suppressors of sensitivity to the DNA topoisomerase I poison camptothecin or the poly(ADP-ribose) polymerase inhibitor olaparib, we detail an approach allowing systematic, large-scale detection of spontaneous or chemically-induced suppressor mutations in yeast or haploid mammalian cells in a short timeframe, and with potential applications in other haploid systems. In addition to applications in molecular biology research, this protocol can be used to identify drug targets and predict drug-resistance mechanisms. Mapping suppressor mutations on the primary or tertiary structures of Protein suppressor hits provides insights into functionally relevant Protein Domains. Importantly, we show that olaparib resistance is linked to missense mutations in the DNA binding regions of PARP1, but not in its catalytic domain. This provides experimental support to the concept of PARP1 trapping on DNA as the prime source of toxicity to PARP inhibitors, and points to a novel olaparib resistance mechanism with potential therapeutic implications.
-
detection of functional Protein Domains by unbiased genome wide forward genetic screening
bioRxiv, 2017Co-Authors: Mareike Herzog, Fabio Puddu, Julia Coates, Nicola J Geisler, Josep V Forment, Stephen P JacksonAbstract:Genetic and chemo-genetic interactions have played key roles in elucidating the molecular mechanisms by which certain chemicals perturb cellular functions. Many studies have employed gene knockout collections or gene disruption/depletion strategies to identify routes for evolving resistance to chemical agents. By contrast, searching for point-mutational genetic suppressors that can identify separation- or gain-of-function mutations, has been limited even in simpler, genetically amenable organisms such as yeast, and has not until recently been possible in mammalian cell culture systems. Here, by demonstrating its utility in identifying suppressors of cellular sensitivity to the drugs camptothecin or olaparib, we describe an approach allowing systematic, large-scale detection of spontaneous or chemically-induced suppressor mutations in yeast and in haploid mouse embryonic stem cells in a short timeframe, and with potential applications in essentially any other haploid system. In addition to its utility for molecular biology research, this protocol can be used to identify drug targets and to predict mechanisms leading to drug resistance. Mapping suppressor mutations on the primary sequence or three-dimensional structures of Protein suppressor hits provides insights into functionally relevant Protein Domains, advancing our molecular understanding of Protein functions, and potentially helping to improve drug design and applicability.
Mareike Herzog - One of the best experts on this subject based on the ideXlab platform.
-
detection of functional Protein Domains by unbiased genome wide forward genetic screening
Scientific Reports, 2018Co-Authors: Mareike Herzog, Fabio Puddu, Julia Coates, Nicola J Geisler, Josep V Forment, Stephen P JacksonAbstract:Establishing genetic and chemo-genetic interactions has played key roles in elucidating mechanisms by which certain chemicals perturb cellular functions. In contrast to gene disruption/depletion strategies to identify mechanisms of drug resistance, searching for point-mutational genetic suppressors that can identify separation- or gain-of-function mutations has been limited. Here, by demonstrating its utility in identifying chemical-genetic suppressors of sensitivity to the DNA topoisomerase I poison camptothecin or the poly(ADP-ribose) polymerase inhibitor olaparib, we detail an approach allowing systematic, large-scale detection of spontaneous or chemically-induced suppressor mutations in yeast or haploid mammalian cells in a short timeframe, and with potential applications in other haploid systems. In addition to applications in molecular biology research, this protocol can be used to identify drug targets and predict drug-resistance mechanisms. Mapping suppressor mutations on the primary or tertiary structures of Protein suppressor hits provides insights into functionally relevant Protein Domains. Importantly, we show that olaparib resistance is linked to missense mutations in the DNA binding regions of PARP1, but not in its catalytic domain. This provides experimental support to the concept of PARP1 trapping on DNA as the prime source of toxicity to PARP inhibitors, and points to a novel olaparib resistance mechanism with potential therapeutic implications.
-
detection of functional Protein Domains by unbiased genome wide forward genetic screening
bioRxiv, 2017Co-Authors: Mareike Herzog, Fabio Puddu, Julia Coates, Nicola J Geisler, Josep V Forment, Stephen P JacksonAbstract:Genetic and chemo-genetic interactions have played key roles in elucidating the molecular mechanisms by which certain chemicals perturb cellular functions. Many studies have employed gene knockout collections or gene disruption/depletion strategies to identify routes for evolving resistance to chemical agents. By contrast, searching for point-mutational genetic suppressors that can identify separation- or gain-of-function mutations, has been limited even in simpler, genetically amenable organisms such as yeast, and has not until recently been possible in mammalian cell culture systems. Here, by demonstrating its utility in identifying suppressors of cellular sensitivity to the drugs camptothecin or olaparib, we describe an approach allowing systematic, large-scale detection of spontaneous or chemically-induced suppressor mutations in yeast and in haploid mouse embryonic stem cells in a short timeframe, and with potential applications in essentially any other haploid system. In addition to its utility for molecular biology research, this protocol can be used to identify drug targets and to predict mechanisms leading to drug resistance. Mapping suppressor mutations on the primary sequence or three-dimensional structures of Protein suppressor hits provides insights into functionally relevant Protein Domains, advancing our molecular understanding of Protein functions, and potentially helping to improve drug design and applicability.
Per Jemth - One of the best experts on this subject based on the ideXlab platform.
-
the transition state structure for coupled binding and folding of disordered Protein Domains
Scientific Reports, 2013Co-Authors: Jakob Dogan, Xin Mu, Åke Engström, Per JemthAbstract:Intrinsically disordered Proteins are abundant in the eukaryotic proteome, and they are implicated in a range of different diseases. However, there is a paucity of experimental data on molecular details of the coupled binding and folding of such Proteins. Two interacting and relatively well studied disordered Protein Domains are the activation domain from the p160 transcriptional co-activator ACTR and the nuclear co-activator binding domain (NCBD) of CREB binding Protein. We have analyzed the transition state for their coupled binding and folding by Protein engineering and kinetic experiments (Φ-value analysis) and found that it involves weak native interactions between the N-terminal helices of ACTR and NCBD, but is otherwise "disordered-like". Most native hydrophobic interactions in the interface between the two Domains form later, after the rate-limiting barrier for association. Linear free energy relationships suggest a cooperative formation of native interactions, reminiscent of the nucleation-condensation mechanism in Protein folding.
-
fast association and slow transitions in the interaction between two intrinsically disordered Protein Domains
Journal of Biological Chemistry, 2012Co-Authors: Jakob Dogan, Åke Engström, Tanja Schmidt, Per JemthAbstract:Proteins that contain long disordered regions are prevalent in the proteome and frequently associated with diseases. However, the mechanisms by which such intrinsically disordered Proteins (IDPs) recognize their targets are not well understood. Here, we report the first experimental investigation of the interaction kinetics of the nuclear co-activator binding domain of CREB-binding Protein and the activation domain from the p160 transcriptional co-activator for thyroid hormone and retinoid receptors. Both Protein Domains are intrinsically disordered in the free state and synergistically fold upon binding each other. Using the stopped-flow technique, we found that the binding reaction is fast, with an association rate constant of 3 × 107 m−1 s−1 at 277 K. Mutation of a conserved buried intermolecular salt bridge showed that electrostatics govern the rapid association. Furthermore, upon mutation of the salt bridge or at high salt concentration, an additional kinetic phase was detected (∼20 and ∼40 s−1, respectively, at 277 K), suggesting that the salt bridge may steer formation of the productive bimolecular complex in an intramolecular step. Finally, we directly measured slow kinetics for the IDP Domains (∼1 s−1 at 277 K) related to conformational transitions upon binding. Together, the experiments demonstrate that the interaction involves several steps and accumulation of intermediate states. Our data are consistent with an induced fit mechanism, in agreement with previous simulations. We propose that the slow transitions may be a consequence of the multipartner interactions of IDPs.
