Introduction:The gene regulation, cell proliferation and the production

Introduction:The Literature I am reviewing refers to the structure of the isoprenylcysteine carboxyl methyltransferase (ICMT) enzyme and how implementing different mechanisms of inhibition will ultimately result in the enzyme’s inactivation. Inhibition of this enzyme is particularly important in the RAS signal transduction pathway as ICMT is a necessity for RAS proteins because it handles the maturation of the RAS GTPases so they can function in gene regulation and cell proliferation. The link between this enzyme and RAS G proteins is oncogenesis. Oncogenesis is the term coined for the development of cancer within the somatic cells (Lodish, H. (2008). Molecular Cell Biology. 6th ed. W. H. Freeman, 2008, p.935.).  RAS proteins are oncogenes which means that they are proteins encoded by DNA that are at a significantly higher risk of becoming cancerous cells. Due to RAS GTPases having a direct link to gene regulation, cell proliferation and the production of second messenger systems, they are an ideal target to aid in cancer metastasis. If mutated, the cancer retains the ability to keep the RAS proteins in their active Guanine nucleotide Exchange Factor bound state. Once the RAS is constantly active the cancer utilises the open pathway to manipulate cancer cell proliferation, promoting the growth of these cells and the spread of the cancer through the blood stream to other parts of the body (Lodish, H. (2008). Molecular Cell Biology. 6th ed. W. H. Freeman, 2008, pp.936-937). Ergo by inhibiting the intramembranous enzyme ICMT, it won’t mature the RAS protein into its active form and as a result cancer cannot manipulate this pathway to induce further development. The article I’m reviewing investigates the crystalline structure of ICMT in the endoplasmic reticulum (ER) by utilising different methods to show how the enzyme can be destabilised and thus aiding in the prevention of RAS- driven cancers and prelamin A progeria (Diver, M., Pedi, L., Koide, A., Koide, S. and Long, S. (2018). Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT. Nature, 553(7689), pp.526-529.).Structure of ICMT:The structure of ICMT in the intramembrane of the ER is essential as the purpose of the paper is to deliver a detailed image of the structure which can be further investigated. This paper is a follow on from another article produced by MM. Diver et al in 2014 where the structure of the enzyme was manipulated to induce inhibition by mutations of essential amino acid (AA) residues within the structure (Diver, M. and Long, S. (2014). Mutational Analysis of the Integral Membrane Methyltransferase Isoprenylcysteine Carboxyl Methyltransferase (ICMT) Reveals Potential Substrate Binding Sites. Journal of Biological Chemistry, 289(38), pp.26007-26020.). The enzyme’s structure is a class four integral protein with eight alpha-helical domains called M1 – M8 that rests in a typical phospholipid bilayer. The structure retains a short cytosolic N-Terminus and a long cytosolic C-Terminus. The terminals refer to the backbone structure of a protein where the N-Terminus stands for an NH3 (amino) region and the C-Terminus stands for a COOH (carboxylic acid) region. The alpha-helices are interconnected by short proline rich residues in the luminal leaflet, which is typical for large membranous proteins for stabilisation. In addition, the structure contains multiple stabilising AA residues, that form hydrogen bonds (H) to ensure total security to the overall structure, specifically to the active site where the environment is rich in aromatic AA residues (Yang, J. et al (2011). Mechanism of Isoprenylcysteine Carboxyl Methylation from the Crystal Structure of the Integral Membrane Methyltransferase ICMT. Molecular Cell, 44(6), pp.997-1004). The helical domains M1 and M2 share non – covalent interactions with the active site in conjunction with the enzyme’s cofactor, further attributing to the powerful stabilising forces. A cofactor or coenzyme is either an essential organic molecule or inorganic ion that is responsible for allowing the function of ICMT (McMurry, J. (2018). Organic Chemistry. 9th ed. Cengage Learning, p.898.).  The coenzyme for ICMT is known as S-adenosyl-Lhomocysteine (AdoHcy) and can only be oriented in the cofactor pocket when its released and S-adenosyl methionine is bound instead which is mediated by hinge point residues, allowing for flexibility in the enzyme.The cofactor pocket is structurally secure by interactions with the M6 and capping protein on the C-terminus. It’s a requirement for the enzyme to facilitate the activation of catalysis, therefore it has stabilising regions in which the active site is associated with. The active site itself has links to M1 and M2 and consists of many hydrophobic AA and polar AA.Figure 1.1: ICMT two-dimensional structure.This figure is a two-dimensional rendition of the ICMT structure. It outlines the enzymes general structure,indicating the positions of the helical domains, terminals, stabilising residues and proline motifs. Italso conveys the position of the active site, substrate binding regions and its associated stabilising residues. Source: Diver, M. and Long, S. (2014). Mutational Analysis of the Integral Membrane Methyltransferase Isoprenylcysteine Carboxyl Methyltransferase (ICMT) Reveals Potential Substrate Binding Sites. Journal of Biological Chemistry, 289(38), pp.26007-26020.Figure 1.2: Hydrogen bondsThis figure outlines the H found between the AA residues of the ICMT structure. The dashed lines denote H.Source: (Yang, J. et al (2011). Mechanism of Isoprenylcysteine Carboxyl Methylation from the Crystal Structure of the Integral Membrane Methyltransferase ICMT. Molecular Cell, 44(6), pp.997-1004).  Figure 1.3: The active site The active site of the enzyme outlined here in three – dimensional form shows the residues that stabilise the active site. Here isthe AdoHcy which is an essential AA derivative. Source: (Yang, J. et al (2011). Mechanism of Isoprenylcysteine Carboxyl Methylation from the Crystal Structure of the Integral Membrane Methyltransferase ICMT. Molecular Cell, 44(6), pp.997-1004). Protein Substrates and RAS Maturation:ICMT’s functions are utilised for the maturation of RAS G proteins and prelamin A. ICMT is responsible for maturing proteins and also acts on more than two hundred other intercellular proteins. However, ICMT acts upon specific proteins known as CAAX proteins. CAAX proteins must undertake three cellular processes but only one is specific to ICMT. ICMT ensures complete maturation of the protein, steps include: polyisoprenylation, proteolysis and carboxyl methylation (Gao L, Liao J, Yang GY. CAAX-box protein, prenylation process and carcinogenesis. Am J Transl Res. 2009;1:312–PMC free article PubMed). The first two steps are vital to the addition of prenyl groups and removal of the -AAX terminal of the CAAX protein but, the final most pivotal step is where ICMT is solely responsible for the methylation of its prenylcysteine substrates, thus maturing it’s protein substrates. The reason ICMT would be found in the ER is due to RAS being sent to the plasma membrane in a secretory vesicle. RAS must be matured by ICMT prior shipping to the cell membrane. Once matured the Ras GTPases are essential to many signal transduction pathways especially Receptor Tyrosine Kinase pathways (RTK) (Gao L, Liao J, Yang GY. CAAX-box protein, prenylation process and carcinogenesis. Am J Transl Res. 2009;1:312–PMC free article PubMed). In this pathway the RAS proteins are activated downstream of the transmembrane receptor. Once a molecule such as Epidermal Growth Factor binds to the RTK, the receptor dimerises and auto-phosphorylates. The phosphate molecules on the cytosolic leaflet of the receptor bind the SH2 and the SH3 (GRB2) proteins which in turn bind the Sos protein. The Sos protein facilitates the conversion of RAS -GAP to RAS -GEF. Once in the active GEF bound state the RAS protein can cause an autophosphorylation cascade via other proteins until specific ones bind to DNA to regulate gene expression. Other proteins include: RAF, ERK, MAP kinases.  RAS proteins are explicitly limited to the plasma membrane of the cell and don’t have locomotion around the cytosol (Wright, L. and Philips, M. (2006). Thematic review series: Lipid Posttranslational ModificationsCAAX modification and membrane targeting of Ras. Journal of Lipid Research, 47(5), pp.883-891.). If a RAS GTPase became permanently active by a mutation then gene regulation would be constantly active, because of this cancer can manipulate the RAS signal transduction pathway to induce cancer cell proliferation and further metastasis within the body. This can make cancers more aggressive and more difficult to cure. This is why proteins linked to ICMT are typically proto-oncogenes.Figure 2.1: RAS Structure This diagram outlines the structure of a matured RAS G protein in both its GEF and GAP forms. The diagram was obtained in the same way as ICMT in Figure 1.2 which is known as X-Ray crystallography. It’s a technique used to figure out the structure of a crystal Source: Lodish, H. (2008). Molecular Cell Biology. 6th ed. W. H. Freeman, 2008, p.542.Figure 2.2: RAS Pathways Here the RAS signal transduction pathway is projecting from the centre conveying its wide reach. Notice must be paid to MEKK, JNK, RAF and ERK as these are the typical proteins manipulated in the cancers rigid progression.Source: Malumbres, M. and Barbacid, M. (2003). Timeline: RAS oncogenes: the first 30 years. Nature Reviews Cancer, 3(6), pp.459-465. Figure 2.3: Cancer FormationShown are RAS proteins becoming cancer cells. As a proto-oncogene, it’s readily manipulated and mutated. In addition to this, the “guardian of the Genome” p53 is a protein that’s responsible for surpressing tumours (Solomon, H., Madar, S. and Rotter, V. (2011). Mutant p53 gain of function is interwoven into the hallmarks of cancer. The Journal of Pathology, 225(4), pp.475-478.). If these two cells are mutated then cancer is an inevitability.Source: Luo, J. and Elledge, S. (2008). Deconstructing   oncogenesis. Nature, 453(7198), pp.995-996.ICMT and OncogenesisICMT is critical to almost all metabolic pathways connected to RAS signal transduction. With RAS being interconnected to ionotropic and metabotropic cell surface receptors it’s a downstream contact point. As aforementioned this makes the RAS protein a target for mutation to cancer and gives it the ability to become defiant. It gives cancer an advantage against host defences and clinical therapies because mutations in specialised proteins allows access in and out of other pathways and for more mutating downstream, avoiding the dependency upon one protein. The genome consists of approximately twenty-five thousand genes and of that number there are roughly twenty-one thousand proteins that are or can be expressed by humans. RAS has some isoforms, but they don’t add up to a significant number within the proteome. Therefore, mutations and deregulations are quite understandable in this context and it becomes increasingly evident as to why RAS proteins create aggressive cancers (Wang, T., et al. (2017). Gene Essentiality Profiling Reveals Gene Networks and Synthetic Lethal Interactions with Oncogenic Ras. Cell, 168(5), pp.890-903.e15.).Figure 3: RAS CancerThis image shows us a benign RAS mutation that is pre-operation. There is a high risk of the mutation becoming malignant.Source: Guttler, D. (2018). RAS Mutation in “benign”( NIFT-P)Non invasive follicular tumor with papillary cell features – Thyroid Center of Santa Monica. online Thyroidnosurgery.com. Available at: http://thyroidnosurgery.com/thyroid-cancer/ras-mutation-in-benign-nift-pnon-invasive-follicular-tumor-with-papillary-cell-features/ Accessed 30 Jan. 2018.ICMT and Progeria: Mutations in the substrates of ICMT aren’t solely linked to GTPases. Prelamin A is another typical protein that matures under methylation. The substrates linked to ICMT converge towards being disease related as does prelamin A. This is a nuclear associated, class five intermediate filament. It’s fibrous in nature and provides crucial support to the regulation of transcription. Its needed role in the cell leads to an association with progeria. Progeroid diseases include, Hutchinson-Gilford progeria syndrome and restrictive dermopathy and both of these arise because of its association with ICMT. Progeria is a fatal condition distinguished by rapid ageing in children. Most children don’t survive to their adolescence and frequently die of heart disease (Ibrahim, M., et al. (2013). Targeting Isoprenylcysteine Methylation Ameliorates Disease in a Mouse Model of Progeria. Science, 340(6138), pp.1330-1333.).Figure 4: Mouse Model of ProgeriaThis is a mouse model of the effects of Progeria. As evident on the left the mouse is aged and deformed. There is a clear issue with its joints as well as improper growth of the body and hair.Source: Johnson, T. (2013). Rapid Aging Rescue?. Science, 340(6138), pp.1299-1300.ICMT Inhibition:  Inhibition of ICMT has great potential for combatting aggressive diseases through various inhibitory modes; structural mutations, monobody insertion and nucleophilic attack. By inhibiting the linking agent between Ras-cancers and prelamin A-progeria, ICMT won’t be able to cause maturation which might be useful for novel clinical treatments. Monobody are synthetic binding proteins that are created using randomised fibronectin protein domains. These are molecules that can be substituted for antibodies in a complex. Considering monobodies are classed as a mimetic, it would insert into the crevice of the ICMT protein domain. There, the monobody’s synthetic residues would interact with ICMT residues, forming H with the residues between the spaces of M1-M8. It’s location prevents the enzyme from interacting with its protein substrates as depicted by X-Ray crystallography (Diver, M. (2018). Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT. Nature, online 553(7689), p.1. Available at: https://www.nature.com/articles/nature25439 Accessed 31 Jan. 2018.). The substrates specificity is due to its positioning and shape but when exposed to the monobody loses it all, owing to the H disrupting the alpha-helices and the monobodies binding next to the active site. Nucleophilicity: A nucleophile is a substance that is nucleus-loving and attacks an electrophile which is electron-loving. In terms of ICMT inhibition, it would be inhibiting the enzyme as multiple residues are running throughout the structure. Using an electron rich atom that is seeking a bond to reach the octet, it’s feasible to inhibit the enzyme, destabilising the alpha-helical domains via nucleophilic attack on electrophilic AA residues (McMurry, J. (2018). Organic Chemistry. 9th ed. Cengage Learning, p.898.).Structural Mutations: ICMT is rigorous because of its eight alpha-helical domains. If certain helices were affected, the enzyme will lose all of its function. If helical domains M1 and M2 were genetically deleted the enzyme is rendered inactive. M4 and M5, when mutated through changes to their AA composition both lose complete functionality. By destabilising the helicases proline rich connectors, they lose all connectivity to the overall structure. These important features directly tie in with the active site and the cofactor pocket. The active site utilises most of the alpha-helices to stabilise its own structure and any mutations that affect them will inhibit it. Therefore, there are two modes of inhibition here, 1) the structure is destabilised and 2) the active site loses its ability to catalyse. The cofactor AdoHcy and its modelled counterpart AdoMet, if mutated to alanine it leaves ICMT with over ninety-five percent inactivity. The cofactor is essential to enzymic function so when it loses functionality the enzyme can’t catalyse reactions. Figure 5.1: MonobodyThis figure outlines the structure of a monobody. Notice the difference between light and heavy chains including the brown structure which is called a FG loop. This is one of the key aspects of the monobody as it plays a role in interacting with ICMT to inhibit it.Source: En.wikipedia.org. (2018). Monobody. online Available at: https://en.wikipedia.org/wiki/Monobody#/media/File:PDB_2rhe_EBI.jpg Accessed 30 Jan. 2018.Figure 5.2: Nucleophilic AttackThis is a generic nucleophilic attack. The Nu denotes the nucleophile which is electron rich as seen by the electrons on the outside. It’s attacking the C atom which only has three bonds out of its required four showing its electron deficient. Once attacked the C shares the electrons with the Nu. Source: Evenson, R. (2018). What is a nucleophilic attack?. online Quora. Available at: https://www.quora.com/What-is-a-nucleophilic-attack Accessed 30 Jan. 2018.Summary/Conclusion:  To summarise, the major findings of this article was the atomic structure of one of the most important enzymes that’s linked to cancers and progeria. X-Ray crystallography was the main technique used to determine the structure/function relationship between mutation and inhibition. These results are novel as it’s the clearest crystallised image of ICMT to date. This idea can be of help in the long-term fight against aggressive diseases as seen in Discover where they look at mutations and genes in cancer therapies (Swartz, A. (2016). Fighting Cancer With Data. Discover, online (October 2016). Available at: http://discovermagazine.com/2016/oct/fighting-cancer-with-data Accessed 30 Jan. 2018.). A combination of these novel ideas might give the necessary knowledge to aid in clinical tests and trials. The next step for this article is to accurately test how inhibition affects ICMT.References1. Lodish, H. (2008). Molecular Cell Biology. 6th ed. W. H. Freeman, 2008, p.935.2. Lodish, H. (2008). Molecular Cell Biology. 6th ed. W. H. Freeman, 2008, pp.936-937.3. Diver, M. (2018). Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT. Nature, online 553(7689), p.1. Available at: https://www.nature.com/articles/nature25439 Accessed 26 Jan. 2018.4. Diver, M. and Long, S. (2014). Mutational Analysis of the Integral Membrane Methyltransferase Isoprenylcysteine Carboxyl Methyltransferase (ICMT) Reveals Potential Substrate Binding Sites. Journal of Biological Chemistry, 289(38), pp.26007-26020.5. Yang, J. (2011). Mechanism of Isoprenylcysteine Carboxyl Methylation from the Crystal Structure of the Integral Membrane Methyltransferase ICMT. Molecular Cell, 44(6), pp.997-1004.6. McMurry, J. (2018). Organic Chemistry. 9th ed. Cengage Learning, p.898.7. Gao L, Liao J, Yang GY. CAAX-box protein, prenylation process and carcinogenesis. Am J Transl Res. 2009;1:312–PMC free article PubMed8. Wright, L. and Philips, M. (2006). Thematic review series: Lipid Posttranslational ModificationsCAAX modification and membrane targeting of Ras. Journal of Lipid Research, 47(5), pp.883-891.9. Lodish, H. (2008). Molecular Cell Biology. 6th ed. W. H. Freeman, 2008, p.542.10. Malumbres, M. and Barbacid, M. (2003). Timeline: RAS oncogenes: the first 30 years. Nature Reviews Cancer, 3(6), pp.459-465. 11. Solomon, H., Madar, S. and Rotter, V. (2011). Mutant p53 gain of function is interwoven into the hallmarks of cancer. The Journal of Pathology, 225(4), pp.475-478.12. Luo, J. and Elledge, S. (2008). Deconstructing oncogenesis. Nature, 453(7198), pp.995-996.13. Wang, T., et al. (2017). Gene Essentiality Profiling Reveals Gene Networks and Synthetic Lethal Interactions with Oncogenic Ras. Cell, 168(5), pp.890-903.e15.14. Guttler, D. (2018). RAS Mutation in “benign”( NIFT-P)Non invasive follicular tumor with papillary cell features – Thyroid Center of Santa Monica. online Thyroidnosurgery.com. Available at: http://thyroidnosurgery.com/thyroid-cancer/ras-mutation-in-benign-nift-pnon-invasive-follicular-tumor-with-papillary-cell-features/ Accessed 30 Jan. 2018.15. Ibrahim, M., et al. (2013). Targeting Isoprenylcysteine Methylation Ameliorates Disease in a Mouse Model of Progeria. Science, 340(6138), pp.1330-1333.16. Johnson, T. (2013). Rapid Aging Rescue?. Science, 340(6138), pp.1299-1300.17. McMurry, J. (2018). Organic Chemistry. 9th ed. Cengage Learning, p.157.18. En.wikipedia.org. (2018). Monobody. online Available at: https://en.wikipedia.org/wiki/Monobody#/media/File:PDB_2rhe_EBI.jpg Accessed 30 Jan. 2018.19. Evenson, R. (2018). What is a nucleophilic attack?. online Quora. Available at: https://www.quora.com/What-is-a-nucleophilic-attack Accessed 30 Jan. 2018.20. Swartz, A. (2016). Fighting Cancer With Data. Discover, online (October 2016). Available at: http://discovermagazine.com/2016/oct/fighting-cancer-with-data Accessed 30 Jan. 2018.

x

Hi!
I'm James!

Would you like to get a custom essay? How about receiving a customized one?

Check it out