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E2 Ubiquitin Conjugating Enzymes
E2 Ubiquitin Conjugating Enzymes
The post-translational modification of proteins with ubiquitin, called ubiquitination, regulates virtually all cellular processes and its perturbations are associated with a range of diseases (Petroski, 2008). Ubiquitin-conjugating enzymes (E2s) act at the core of the ubiquitination cascade: they transfer the activated ubiquitin from an E1 (ubiquitin-activating enzyme) to the substrate, usually with assistance of an E3 ligase, and are responsible for determining the typology and extent of modification (van Wijk and Timmers, 2010). To accomplish their function, the 38 human E2s interact with several components of the ubiquitination cascade with varying degree of specificity and are indispensable for the ubiquitination reactions to take place. Consequently, defects in E2 activities have been associated with human diseases such as cancer, neurodegenerative and immune disorders. However, only 9 chemical modulators of the E2 enzyme activities have been reported and in most cases they are unspecific and/or of low potency (Liu et al., 2015). In the Auer lab, we have produced 13 active human E2s, most of which have already been associated with various human diseases. These enzymes, expressed and purified as fluorescent protein fusions, screened in our proprietary ubiquitination assay, which is currently in the PCT stage.
The post-translational modification of proteins with ubiquitin, called ubiquitination, regulates virtually all cellular processes and its perturbations are associated with a range of diseases (Petroski, 2008). Ubiquitin-conjugating enzymes (E2s) act at the core of the ubiquitination cascade: they transfer the activated ubiquitin from an E1 (ubiquitin-activating enzyme) to the substrate, usually with assistance of an E3 ligase, and are responsible for determining the typology and extent of modification (van Wijk and Timmers, 2010). To accomplish their function, the 38 human E2s interact with several components of the ubiquitination cascade with varying degree of specificity and are indispensable for the ubiquitination reactions to take place. Consequently, defects in E2 activities have been associated with human diseases such as cancer, neurodegenerative and immune disorders. However, only 9 chemical modulators of the E2 enzyme activities have been reported and in most cases they are unspecific and/or of low potency (Liu et al., 2015). In the Auer lab, we have produced 13 active human E2s, most of which have already been associated with various human diseases. These enzymes, expressed and purified as fluorescent protein fusions, screened in our proprietary ubiquitination assay, which is currently in the PCT stage.
[1] Petroski, M.D. (2008). The ubiquitin system, disease, and drug discovery. BMC Biochem. 9 Suppl 1, S7.
[1] Petroski, M.D. (2008). The ubiquitin system, disease, and drug discovery. BMC Biochem. 9 Suppl 1, S7.
[2] van Wijk, S.J.L., and Timmers, H.T.M. (2010). The family of ubiquitin-conjugating enzymes (E2s): deciding between life and death of proteins. FASEB J. 24, 981–993.
[2] van Wijk, S.J.L., and Timmers, H.T.M. (2010). The family of ubiquitin-conjugating enzymes (E2s): deciding between life and death of proteins. FASEB J. 24, 981–993.
[3] Liu, J., Shaik, S., Dai, X., Wu, Q., Zhou, X., Wang, Z., and Wei, W. (2015). Targeting the ubiquitin pathway for cancer treatment. Biochim. Biophys. Acta 1855, 50–60.
[3] Liu, J., Shaik, S., Dai, X., Wu, Q., Zhou, X., Wang, Z., and Wei, W. (2015). Targeting the ubiquitin pathway for cancer treatment. Biochim. Biophys. Acta 1855, 50–60.
The Ska complex: a key regulator of cell division as a novel target in cancer therapy
The Ska complex: a key regulator of cell division as a novel target in cancer therapy
Cell division is a crucial process of life which assures precise transfer of the genetic material of a cell by equally distributing the duplicated chromosomes to its daughter cells. Proper physical attachment of centromeric chromatin to spindle microtubules mediated by kinetochores is a primary requirement for correct chromosome alignment and segregation. A key component involved in this process is a three subunit protein complex called the Ska complex, comprising Ska1, Ska2 and Ska3 [1,2,3]. Depletion of this complex from eukaryotic somatic cells induces defects in chromosome segregation and increased cell death [1,2,4]. Work from different laboratories have shown that the Ska complex exerts its function mainly by stabilizing kinetochore-microtubule connections by directly binding along polymerized microtubules [2,3,5,6]. Within the Ska complex, the C-terminal domain of Ska1 - referred to as Ska1-MTBD (Micro Tubule Binding Domain) - is the main microtubule-binding element. The structure and microtubule binding properties of the Ska1-MTBD have been recently characterized in Jeyaprakash Arulanandam’s lab. Ska1-MTBD contains a modified winged-helix motif which is common in nucleic acid binding proteins. Structural data together with crosslinking/MS (CLMS) data allowed the identification of surface patches crucial for its microtubule binding ability [6]. Ska1 has been shown to be overexpressed in various cancers, including gastric and oral. It has also been suggested that inhibiting Ska1 could be a novel approach in cancer therapy [7,8,9,10]. Having the reagents and right tools necessary to investigate the Ska1-MTBD we are in a great position to explore the possibilities of targeting Ska1-MTBD. - Jeyaprakash Arulanandam
Cell division is a crucial process of life which assures precise transfer of the genetic material of a cell by equally distributing the duplicated chromosomes to its daughter cells. Proper physical attachment of centromeric chromatin to spindle microtubules mediated by kinetochores is a primary requirement for correct chromosome alignment and segregation. A key component involved in this process is a three subunit protein complex called the Ska complex, comprising Ska1, Ska2 and Ska3 [1,2,3]. Depletion of this complex from eukaryotic somatic cells induces defects in chromosome segregation and increased cell death [1,2,4]. Work from different laboratories have shown that the Ska complex exerts its function mainly by stabilizing kinetochore-microtubule connections by directly binding along polymerized microtubules [2,3,5,6]. Within the Ska complex, the C-terminal domain of Ska1 - referred to as Ska1-MTBD (Micro Tubule Binding Domain) - is the main microtubule-binding element. The structure and microtubule binding properties of the Ska1-MTBD have been recently characterized in Jeyaprakash Arulanandam’s lab. Ska1-MTBD contains a modified winged-helix motif which is common in nucleic acid binding proteins. Structural data together with crosslinking/MS (CLMS) data allowed the identification of surface patches crucial for its microtubule binding ability [6]. Ska1 has been shown to be overexpressed in various cancers, including gastric and oral. It has also been suggested that inhibiting Ska1 could be a novel approach in cancer therapy [7,8,9,10]. Having the reagents and right tools necessary to investigate the Ska1-MTBD we are in a great position to explore the possibilities of targeting Ska1-MTBD. - Jeyaprakash Arulanandam
[1] Gaitaons et al., EMBO J (2009) 28(10): 1442-52.
[1] Gaitaons et al., EMBO J (2009) 28(10): 1442-52.
[2] Welburn et al., Dev Cell (2009) 16(3): 374-85.
[2] Welburn et al., Dev Cell (2009) 16(3): 374-85.
[3] Jeyaprakash et al., Mol Cell (2012) 46(3): 274-86.
[3] Jeyaprakash et al., Mol Cell (2012) 46(3): 274-86.
[4] Daum et al., Curr Biol (2009) 19(17): 1467-72.
[4] Daum et al., Curr Biol (2009) 19(17): 1467-72.
[5] Schmidt et al., Dev Cell (2012) 23(5): 968-80.
[5] Schmidt et al., Dev Cell (2012) 23(5): 968-80.
[6] Abad et al., Nat Commun. (2014) 5:2964.
[6] Abad et al., Nat Commun. (2014) 5:2964.
[7] Zhang et al., Cancer Cell Int. (2013) 13(1): 83.
[7] Zhang et al., Cancer Cell Int. (2013) 13(1): 83.
[8] Sun et al., Mol Cell Biochem (2014) 391(1-2): 167-74.
[8] Sun et al., Mol Cell Biochem (2014) 391(1-2): 167-74.
[9] Li et al., J. Pathol (2014) 234(2): 178-89.
[9] Li et al., J. Pathol (2014) 234(2): 178-89.
[10] Shi et al., Mol. Med. Rep (2015) 11(5): 3533-8.
[10] Shi et al., Mol. Med. Rep (2015) 11(5): 3533-8.
Alpha-Synuclein: A major component in Lewy bodies a pathological hallmarks of Parkinson’s Disease.
Alpha-Synuclein: A major component in Lewy bodies a pathological hallmarks of Parkinson’s Disease.
Alpha-Synuclein (αS) is a protein which has been shown to be a major component in Lewy bodies (LB). Lewy bodies are filamentous aggregates that are a pathological hallmarks of Parkinson’s Disease (PD) and other synucleopathies. αS is an intrinsically unfolded protein that aggregates in pathological conditions, first it aggregates forming oligomers, that are thought to be the toxic species, and then forming the fibrils that are present in LB. Many small molecules and peptides have been shown to inhibit alpha-synuclein fibrillation. The principal assay that has been used to study this is based on the fluorescence dye Thioflavin T, which binds to fibrils. Our goal in this project is to develop novel miniaturised assays to monitor the earliest steps of alpha-Synuclein oligomerization using CTB's micro-bead / CONA technologies and to apply the assays to identify novel small molecules, fragments, peptidomimetics, miniproteins/biosimilars and natural products as chemical tools to increase our knowledge of the alpha-Synucleins aggregation mechanism and ideally, as starting points for lead discovery
Alpha-Synuclein (αS) is a protein which has been shown to be a major component in Lewy bodies (LB). Lewy bodies are filamentous aggregates that are a pathological hallmarks of Parkinson’s Disease (PD) and other synucleopathies. αS is an intrinsically unfolded protein that aggregates in pathological conditions, first it aggregates forming oligomers, that are thought to be the toxic species, and then forming the fibrils that are present in LB. Many small molecules and peptides have been shown to inhibit alpha-synuclein fibrillation. The principal assay that has been used to study this is based on the fluorescence dye Thioflavin T, which binds to fibrils. Our goal in this project is to develop novel miniaturised assays to monitor the earliest steps of alpha-Synuclein oligomerization using CTB's micro-bead / CONA technologies and to apply the assays to identify novel small molecules, fragments, peptidomimetics, miniproteins/biosimilars and natural products as chemical tools to increase our knowledge of the alpha-Synucleins aggregation mechanism and ideally, as starting points for lead discovery
Survivin: Key regulator of mitosis and apoptosis, a therapeutic target in oncology
Survivin: Key regulator of mitosis and apoptosis, a therapeutic target in oncology
Malignancies are often described in reference to the “hallmarks of cancer”, two of which are resistance to cell death and limitless replicative potential [1].
Malignancies are often described in reference to the “hallmarks of cancer”, two of which are resistance to cell death and limitless replicative potential [1].
Survivin is the smallest member of the inhibitor of apoptosis (IAP) family, consisting of a globular N-terminal domain (known as the baculoviral IAP repeat (BIR)) and a long C-terminal helix [2]. Survivin is upregulated in a range of cancers, including lung, breast, pancreatic and oesophageal cancer among others [3].
Survivin is the smallest member of the inhibitor of apoptosis (IAP) family, consisting of a globular N-terminal domain (known as the baculoviral IAP repeat (BIR)) and a long C-terminal helix [2]. Survivin is upregulated in a range of cancers, including lung, breast, pancreatic and oesophageal cancer among others [3].
This overexpression highlights the importance of survivin in understanding the process of carcinogenesis. Interestingly many studies have shown Survivin to play a role in both the hallmarks of cancer mentioned previously.
This overexpression highlights the importance of survivin in understanding the process of carcinogenesis. Interestingly many studies have shown Survivin to play a role in both the hallmarks of cancer mentioned previously.
Survivin was originally described as an inhibitor of caspase-9, and consequently an inhibitor of apoptosis, although this is now a subject of debate. More recently however Survivin has also been shown to have a key function in the regulation of the mitotic spindle checkpoint and accurate division of genetic material during mitosis [4]. This mitotic function is achieved due to the central role of Survivin in the chromosomal passenger complex (CPC), which consists of three other proteins. Borealin and INCENP interact with the helical domain of survivin to form a three-helical bundle (Figure 1), while aurora B, the enzymatic component binds to INCENP [5].
Survivin was originally described as an inhibitor of caspase-9, and consequently an inhibitor of apoptosis, although this is now a subject of debate. More recently however Survivin has also been shown to have a key function in the regulation of the mitotic spindle checkpoint and accurate division of genetic material during mitosis [4]. This mitotic function is achieved due to the central role of Survivin in the chromosomal passenger complex (CPC), which consists of three other proteins. Borealin and INCENP interact with the helical domain of survivin to form a three-helical bundle (Figure 1), while aurora B, the enzymatic component binds to INCENP [5].
Survivin, along with the proteins it binds in the CPC subsequently plays a critical role in the localisation of aurora B during cell division. The functionality of survivin as both a regulator of cell division and cell death in combination with its overexpression in a wide variety of cancers highlights it as an exciting target in both the understanding and targeting of cancer.
Survivin, along with the proteins it binds in the CPC subsequently plays a critical role in the localisation of aurora B during cell division. The functionality of survivin as both a regulator of cell division and cell death in combination with its overexpression in a wide variety of cancers highlights it as an exciting target in both the understanding and targeting of cancer.
[1] Hanahan, D. and Weinber, R.A. (2011) Hallmarks of cancer: the next generation. Cell. 144 (5), pp. 646-674
[1] Hanahan, D. and Weinber, R.A. (2011) Hallmarks of cancer: the next generation. Cell. 144 (5), pp. 646-674
[2] Jeyaprakash, A.A., Klein, U.R., Lindner, D., Ebert, J., Nigg, E.A. and Conti, E. (2007) Structure of a Survivin-Borealin-INCENP Core Complex Reveals how Chromosomal Passengers Travel Together. Cell. 131 (2), pp. 271-285
[2] Jeyaprakash, A.A., Klein, U.R., Lindner, D., Ebert, J., Nigg, E.A. and Conti, E. (2007) Structure of a Survivin-Borealin-INCENP Core Complex Reveals how Chromosomal Passengers Travel Together. Cell. 131 (2), pp. 271-285
[3] Jaiswal, P.K., Goel, A. and Mittal, R.D. (2015) Survivin: A Molecular Biomarker in Cancer. Indian Journal of Medical Research. 141 (4), pp. 389-397
[3] Jaiswal, P.K., Goel, A. and Mittal, R.D. (2015) Survivin: A Molecular Biomarker in Cancer. Indian Journal of Medical Research. 141 (4), pp. 389-397
[4] Mita, A.C., Mita, M.M., Nawrocki, S.T. and Giles, F.J. (2008) Survivin: Key regulator of mitosis and apoptosis and novel target for cancer therapeutics. Clinical Cancer Research. 14 (16), pp. 5000-5005
[4] Mita, A.C., Mita, M.M., Nawrocki, S.T. and Giles, F.J. (2008) Survivin: Key regulator of mitosis and apoptosis and novel target for cancer therapeutics. Clinical Cancer Research. 14 (16), pp. 5000-5005
[5] Carmena, M., Wheelock, M., Funabiki, H. and Earnshaw, W.C. (2012) The Chromosomal Passenger Complex: From Easy Rider to the Godfather of Mitosis. Nature Reviews Molecular Cell Biology. 13 (12), pp. 789-803
[5] Carmena, M., Wheelock, M., Funabiki, H. and Earnshaw, W.C. (2012) The Chromosomal Passenger Complex: From Easy Rider to the Godfather of Mitosis. Nature Reviews Molecular Cell Biology. 13 (12), pp. 789-803
Figure 1 – Jeyaprakash, A.A., Basquin, C., Jayachandran, U. and Conti, E. (2011) Structural Basis for the Recognition of Phosphorylated Histone H3 by the Survivin Subunit of the Chromosomal Passenger Complex. Cell. 19 (11), pp. 1625-1634
Figure 1 – Jeyaprakash, A.A., Basquin, C., Jayachandran, U. and Conti, E. (2011) Structural Basis for the Recognition of Phosphorylated Histone H3 by the Survivin Subunit of the Chromosomal Passenger Complex. Cell. 19 (11), pp. 1625-1634
Shugoshin: A regulator of localisation/dissociation of the chromosomal passenger complex to the centromere
Shugoshin: A regulator of localisation/dissociation of the chromosomal passenger complex to the centromere
Proper chromosomal segregation is a crucial process in cellular division. Improper segregation can have significant effects on daughter cells, including aneuploidy and cell death. Unsurprisingly defects in this process lead to chromosomal instability (CIN) [1], a key driver of carcinogenesis. This importance is highlighted by a number of proteins responsible for accurate segregation.
Proper chromosomal segregation is a crucial process in cellular division. Improper segregation can have significant effects on daughter cells, including aneuploidy and cell death. Unsurprisingly defects in this process lead to chromosomal instability (CIN) [1], a key driver of carcinogenesis. This importance is highlighted by a number of proteins responsible for accurate segregation.
One such protein is Shugoshin (Japanese for “guardian spirit”). Shugoshin plays key roles in both meiosis and mitosis [2]. During meiotic division Shugoshin is responsible for the protection of centromeric cohesins from seperase during prophase. This function is critical for sister chromatid cohesion [3].
One such protein is Shugoshin (Japanese for “guardian spirit”). Shugoshin plays key roles in both meiosis and mitosis [2]. During meiotic division Shugoshin is responsible for the protection of centromeric cohesins from seperase during prophase. This function is critical for sister chromatid cohesion [3].
Of interest to the Auer lab is shugoshins functionality in mitotic cells. Shugoshin is suggested to be a key component contributing to the timely and proper localisation/dissociation of the chromosomal passenger complex to the centromere [4]. This is due to a tetrapeptide N-terminal sequence (A-K-E-R) that is able to directly bind the baculoviral IAP repeat (BIR) domain of survivin (another key target of the Auer lab) [4].
Of interest to the Auer lab is shugoshins functionality in mitotic cells. Shugoshin is suggested to be a key component contributing to the timely and proper localisation/dissociation of the chromosomal passenger complex to the centromere [4]. This is due to a tetrapeptide N-terminal sequence (A-K-E-R) that is able to directly bind the baculoviral IAP repeat (BIR) domain of survivin (another key target of the Auer lab) [4].
The importance of Shugoshin in cellular division, along with its direct relation to the function of survivin has implications in cancer. This identifies shugoshin as a protein worthy of further study, and an exciting prospect for the Auer lab.
The importance of Shugoshin in cellular division, along with its direct relation to the function of survivin has implications in cancer. This identifies shugoshin as a protein worthy of further study, and an exciting prospect for the Auer lab.
[1] – Thompson, S.L., Bakhoum, S.F., Compton, D.A., (2010) Mechanisms of Chromosomal Instability. Current Biology. 20 (6), pp. 285-295
[1] – Thompson, S.L., Bakhoum, S.F., Compton, D.A., (2010) Mechanisms of Chromosomal Instability. Current Biology. 20 (6), pp. 285-295
[2] – Kiburz, B.M., Amon, A., Marston, A.L., (2008) Shugoshin promotes sister kinetochore biorientation in Saccharomyces cerevisiae. Molecular Biology of the Cell. 19 (3), pp. 1199-1209
[2] – Kiburz, B.M., Amon, A., Marston, A.L., (2008) Shugoshin promotes sister kinetochore biorientation in Saccharomyces cerevisiae. Molecular Biology of the Cell. 19 (3), pp. 1199-1209
[3] – Watanabe, Y., (2005) Shugoshin: Guardian Spirit at the Centromere. Current Opinion in Cell Biology. 17 (6), pp. 590-595
[3] – Watanabe, Y., (2005) Shugoshin: Guardian Spirit at the Centromere. Current Opinion in Cell Biology. 17 (6), pp. 590-595
[4] – Jeyaprakash, A.A., Basquin, C., Jayachandran, U., Conti, E., (2011) Structural basis for the recognition of phosphorylated histone H3 by the survivin subunit of the chromosomal passenger complex. Structure. 19 (11), pp. 1625-1634
[4] – Jeyaprakash, A.A., Basquin, C., Jayachandran, U., Conti, E., (2011) Structural basis for the recognition of phosphorylated histone H3 by the survivin subunit of the chromosomal passenger complex. Structure. 19 (11), pp. 1625-1634
Centromere protein 32 (CENP-32)
Centromere protein 32 (CENP-32)
Centrosomes are essential for spindle formation, spindle pole orientation and correct chromosome segregation in animal cells. During metaphase centromere protein 32 (CENP-32) is required for the association of the centrosome to the spindle pole. Bioinformatic analysis suggests that CENP-32 is a member of the SPOUT family of methyl transferases (MTases), a distinct class of S-Adenosyl-L-Methionine-dependent MTases. CENP-32’s function is predicted to be similar to CLASP1 and CLASP2 which are known to associate with microtubules, centrosomes and kinetochores. They are involved in mitotic spindle assembly and positioning as well as stabilization of microtubule (+)-ends at the cell cortex. CENP-32 is composed of an uncommon alpha/beta fold with a deep topological knot. It is atypical in possessing an additional oligonucleotide-binding fold within the catalytic domain. CENP-32, tagged with streptavidin-binding peptide, shows accumulation on mitotic spindles and kinetochores.
Centrosomes are essential for spindle formation, spindle pole orientation and correct chromosome segregation in animal cells. During metaphase centromere protein 32 (CENP-32) is required for the association of the centrosome to the spindle pole. Bioinformatic analysis suggests that CENP-32 is a member of the SPOUT family of methyl transferases (MTases), a distinct class of S-Adenosyl-L-Methionine-dependent MTases. CENP-32’s function is predicted to be similar to CLASP1 and CLASP2 which are known to associate with microtubules, centrosomes and kinetochores. They are involved in mitotic spindle assembly and positioning as well as stabilization of microtubule (+)-ends at the cell cortex. CENP-32 is composed of an uncommon alpha/beta fold with a deep topological knot. It is atypical in possessing an additional oligonucleotide-binding fold within the catalytic domain. CENP-32, tagged with streptavidin-binding peptide, shows accumulation on mitotic spindles and kinetochores.
Depletion of CENP-32 appears to release centrosomes from bipolar spindles resulting in anastral spindles which appear to function normally in aligning the chromosomes to a metaphase plate and entering anaphase. CENP-32 appears to be required for centrosomes to integrate into fully functional spindles that nucleate astral microtubules as well as nucleate and bind to kinetochore central spindle microtubules. This enzyme, with its critical role in cell division, has not yet been chemically validated or de-validated as a drug target.
Depletion of CENP-32 appears to release centrosomes from bipolar spindles resulting in anastral spindles which appear to function normally in aligning the chromosomes to a metaphase plate and entering anaphase. CENP-32 appears to be required for centrosomes to integrate into fully functional spindles that nucleate astral microtubules as well as nucleate and bind to kinetochore central spindle microtubules. This enzyme, with its critical role in cell division, has not yet been chemically validated or de-validated as a drug target.
1. Maiato H., Khodjakov A., Rieder C.L. 2005. Drosophila CLASP is required for the incorporation of microtubule subunits into fluxing kinetochore fibres. Nat. Cell Biol. 7:42–47
1. Maiato H., Khodjakov A., Rieder C.L. 2005. Drosophila CLASP is required for the incorporation of microtubule subunits into fluxing kinetochore fibres. Nat. Cell Biol. 7:42–47
2. Mimori-Kiyosue Y. et al. (2005) CLASP1 and CLASP2 bind to EB1 and regulate microtubule plus end dynamics at the cell cortex. J Cell Biol. 169(1): 141-53
2. Mimori-Kiyosue Y. et al. (2005) CLASP1 and CLASP2 bind to EB1 and regulate microtubule plus end dynamics at the cell cortex. J Cell Biol. 169(1): 141-53
3. Miller P.M. et al. (2010) Golgi-derived CLASP-dependent Microtubules Control Golgi Organization and Polarized Trafficking in Motile Cells. Nat Cell. Biol. 11(9): 1069-1080
3. Miller P.M. et al. (2010) Golgi-derived CLASP-dependent Microtubules Control Golgi Organization and Polarized Trafficking in Motile Cells. Nat Cell. Biol. 11(9): 1069-1080
4. Reis R. et al. (2009) Dynein and Mast/Orbit/CLASP have antagonistic roles in regulating kinetochore-microtubule plus-end dynamics. J Cell Sci. 122: 2543-2553
4. Reis R. et al. (2009) Dynein and Mast/Orbit/CLASP have antagonistic roles in regulating kinetochore-microtubule plus-end dynamics. J Cell Sci. 122: 2543-2553
5. Patel K. et al. (2012) Multiple Domains of Human CLASP Contribute to Microtubule Dynamics and Organization In Vitro and in Xenopus Egg Extracts. Cytoskeleton. 000: 000-000
5. Patel K. et al. (2012) Multiple Domains of Human CLASP Contribute to Microtubule Dynamics and Organization In Vitro and in Xenopus Egg Extracts. Cytoskeleton. 000: 000-000
6. Bird S.L. et al. (2013) RanGTP and CLASP1 cooperate to position the mitotic spindle. Mol Biol Cell. 24(16): 2506-2514
6. Bird S.L. et al. (2013) RanGTP and CLASP1 cooperate to position the mitotic spindle. Mol Biol Cell. 24(16): 2506-2514
7. Espiritu E.B. et al. (2012) CLASPs function redundantly to regulate astral micorotubules in the C. elegans embryo. Dev Biol. 368(2): 242-254
7. Espiritu E.B. et al. (2012) CLASPs function redundantly to regulate astral micorotubules in the C. elegans embryo. Dev Biol. 368(2): 242-254
8. Kiyomitsu T. and Cheeseman I.M. (2012) Chromosome- an
8. Kiyomitsu T. and Cheeseman I.M. (2012) Chromosome- an
Nus-Complex (targeting protein-protein interactions in small transcription factors)
Nus-Complex (targeting protein-protein interactions in small transcription factors)
PPIs involved in transcription are an attractive option for pharmacological intervention. A host of regulatory proteins are involved in this complex pathway, and disruption of this process has been shown to be of therapeutic benefit. One such PPI important in the regulation of transcription is the interaction between transcription factors N-utilisation substance (Nus) E and NusB (collectively as NusE-NusB). This PPI is critical for formation of the highly regulatory antitermination complex in prokaryotes and to allow stable RNA transcription. Originally discovered in bacteriophage λ, antitermination is unique to prokaryotes, and is the cell’s aid to fix premature termination of RNA synthesis during transcription. Transcription and translation in prokaryotes are directly coupled and transcription antitermination allows the cell to progress from early gene expression to delayed early gene expression. The importance of the NusE-NusB binding interaction was described separately by Robledo (1991), Court (1995) and later by Lou (2008). It was shown that by incorporating point mutations separately into NusE and NusB, this resulted in a reduced protein-protein binding affinity, which subsequently affects the formation of the antitermination complex. The mutations affected the ability of E. coli cells to efficiently transcribe the 16S and 23S ribosomal transcripts, therefore leading to a reduced number of new ribosomes and, hence, a decrease in cell growth.
PPIs involved in transcription are an attractive option for pharmacological intervention. A host of regulatory proteins are involved in this complex pathway, and disruption of this process has been shown to be of therapeutic benefit. One such PPI important in the regulation of transcription is the interaction between transcription factors N-utilisation substance (Nus) E and NusB (collectively as NusE-NusB). This PPI is critical for formation of the highly regulatory antitermination complex in prokaryotes and to allow stable RNA transcription. Originally discovered in bacteriophage λ, antitermination is unique to prokaryotes, and is the cell’s aid to fix premature termination of RNA synthesis during transcription. Transcription and translation in prokaryotes are directly coupled and transcription antitermination allows the cell to progress from early gene expression to delayed early gene expression. The importance of the NusE-NusB binding interaction was described separately by Robledo (1991), Court (1995) and later by Lou (2008). It was shown that by incorporating point mutations separately into NusE and NusB, this resulted in a reduced protein-protein binding affinity, which subsequently affects the formation of the antitermination complex. The mutations affected the ability of E. coli cells to efficiently transcribe the 16S and 23S ribosomal transcripts, therefore leading to a reduced number of new ribosomes and, hence, a decrease in cell growth.
The image at the right shows the structure of the NusE-NusB PPI (PDB ID: 3D3B) NusE: pale orange. NusB: cyan.
The image at the right shows the structure of the NusE-NusB PPI (PDB ID: 3D3B) NusE: pale orange. NusB: cyan.
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