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==Monoglyceride Lipase (MGL)== | ==Monoglyceride Lipase (MGL)== | ||
<StructureSection load='' size='450' side='right' scene='58/580298/Overall_structure/2' caption='Secondary structure of MGL'> | <StructureSection load='' size='450' side='right' scene='58/580298/Overall_structure/2' caption='Secondary structure of MGL'> | ||
[[Image:Complete_crystal_structure.png|left|300px|thumb|Crystal Structure of MGL]] | [[Image:Complete_crystal_structure.png|left|300px|thumb|'''Figure 1:'''Crystal Structure of MGL Alpha helixes are in blue and beta sheets in purple. This protein is a dimer that is linked by antiparallel beta sheets]] | ||
==Background== | ==Background== | ||
Monoglyceride lipase is part of the α/β hydrolase family, having a Ser-His-Asp catalytic triad <ref name="Clemente">[Clemente, J. C., E. Nulton, M. Nelen, M. J. Todd, D. Maguire, C. Schalk-Hihi, L. C. Kuo, S.-P. Zhang, C. M. Flores, and J. K. Kranz. "Screening and Characterization of Human Monoglyceride Lipase Active Site Inhibitors Using Orthogonal Binding and Functional Assays." Journal of Biomolecular Screening 17.5 (2012): 629-40]</ref>. This enzyme is present in most cells, providing the rate limiting step for MG <ref name="Taschler">[Taschler, U., F. P. W. Radner, C. Heier, R. Schreiber, M. Schweiger, G. Schoiswohl, K. Preiss-Landl, D. Jaeger, B. Reiter, H. C. Koefeler, J. Wojciechowski, C. Theussl, J. M. Penninger, A. Lass, G. Haemmerle, R. Zechner, and R. Zimmermann. "Monoglyceride Lipase Deficiency in Mice Impairs Lipolysis and Attenuates Diet-induced Insulin Resistance." Journal of Biological Chemistry 286.20 (2011): 17467-7477]</ref>. MGL terminates the signaling of a primary endocannabinoid, 2-AG <ref name="Savinainen">[Savinainen, Juha R., Megumi Yoshino, Anna Minkkilä, Tapio Nevalainen, and Jarmo T. Laitinen. "Characterization of Binding Properties of Monoglyceride Lipase Inhibitors by a Versatile Fluorescence-based Technique." Analytical Biochemistry 399.1 (2010): 132-34]</ref>MGL is the main enzyme respondsible for hydrolyzing 2-arachidonoylglycerol into arachidonic acid and glycerol ''in vivo'' <ref name="Bertrand">[ Bertrand, T., F. Augé, J. Houtmann, A. Rak, F. Vallée, V. Mikol, P.f. Berne, N. Michot, D. Cheuret, C. Hoornaert, and M. Mathieu. "Structural Basis for Human Monoglyceride Lipase Inhibition." Journal of Molecular Biology 396.3 (2010): 663-73.]</ref>. One of the key features of MGL is the hydrophobic tunnel, which has been suggested to provide a model for drug research. | Monoglyceride lipase is part of the α/β hydrolase family, having a Ser-His-Asp catalytic triad <ref name="Clemente">[Clemente, J. C., E. Nulton, M. Nelen, M. J. Todd, D. Maguire, C. Schalk-Hihi, L. C. Kuo, S.-P. Zhang, C. M. Flores, and J. K. Kranz. "Screening and Characterization of Human Monoglyceride Lipase Active Site Inhibitors Using Orthogonal Binding and Functional Assays." Journal of Biomolecular Screening 17.5 (2012): 629-40]</ref>. This enzyme is present in most cells, providing the rate limiting step for MG <ref name="Taschler">[Taschler, U., F. P. W. Radner, C. Heier, R. Schreiber, M. Schweiger, G. Schoiswohl, K. Preiss-Landl, D. Jaeger, B. Reiter, H. C. Koefeler, J. Wojciechowski, C. Theussl, J. M. Penninger, A. Lass, G. Haemmerle, R. Zechner, and R. Zimmermann. "Monoglyceride Lipase Deficiency in Mice Impairs Lipolysis and Attenuates Diet-induced Insulin Resistance." Journal of Biological Chemistry 286.20 (2011): 17467-7477]</ref>. MGL terminates the signaling of a primary endocannabinoid, 2-AG <ref name="Savinainen">[Savinainen, Juha R., Megumi Yoshino, Anna Minkkilä, Tapio Nevalainen, and Jarmo T. Laitinen. "Characterization of Binding Properties of Monoglyceride Lipase Inhibitors by a Versatile Fluorescence-based Technique." Analytical Biochemistry 399.1 (2010): 132-34]</ref>MGL is the main enzyme respondsible for hydrolyzing 2-arachidonoylglycerol into arachidonic acid and glycerol ''in vivo'' <ref name="Bertrand">[ Bertrand, T., F. Augé, J. Houtmann, A. Rak, F. Vallée, V. Mikol, P.f. Berne, N. Michot, D. Cheuret, C. Hoornaert, and M. Mathieu. "Structural Basis for Human Monoglyceride Lipase Inhibition." Journal of Molecular Biology 396.3 (2010): 663-73.]</ref>. One of the key features of MGL is the hydrophobic tunnel, which has been suggested to provide a model for drug research. | ||
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===Inhibition of MGL=== | ===Inhibition of MGL=== | ||
The importance of inhibiting Monoglyceride lipase is to keep it from breaking down 2-arachidonoyl glycerol. When 2-Ag is broken down it is not able to suppress pain and depression brain functions that human beings experience. N-arachidonyl maleimide (NAM) is one inhibitor of MGL that reacts with the amino acid <scene name='58/580298/Cys252/1'>Cys252</scene>. [[Image:NAM.png]] | The importance of inhibiting Monoglyceride lipase is to keep it from breaking down 2-arachidonoyl glycerol. When 2-Ag is broken down it is not able to suppress pain and depression brain functions that human beings experience. N-arachidonyl maleimide (NAM) is one inhibitor of MGL that reacts with the amino acid <scene name='58/580298/Cys252/1'>Cys252</scene>. [[Image:NAM.png|thumb|'''Figure 2:''' The structure of N-arachidonyl maleimide (NAM)that interacts with Cys252.]] | ||
This Cysteine is buried in the active site near the catalytic serine and functions by sterically clashing with the natural ligand. A possible conformational change to Cys252 upon the binding of NAM could also lead to an inactive form of MGL. | This Cysteine is buried in the active site near the catalytic serine and functions by sterically clashing with the natural ligand. A possible conformational change to Cys252 upon the binding of NAM could also lead to an inactive form of MGL. | ||
MGL is also inhibited by being in complex with <scene name='58/580298/Sar629/2'>SAR629</scene> that is covalently bound to the catalytic Ser132. SAR629 adopts a Y shape and interacts with the MGL by hydrophobic interactions, with a few polar interactions as well. [[Image:SAR.png]] | MGL is also inhibited by being in complex with <scene name='58/580298/Sar629/2'>SAR629</scene> that is covalently bound to the catalytic Ser132. SAR629 adopts a Y shape and interacts with the MGL by hydrophobic interactions, with a few polar interactions as well. [[Image:SAR.png|thumb|'''Figure 3:''' The structure and shape of SAR629.]] | ||
With SAR629 interacting with the catalytic triad it inhibits the triad from breaking down 2-AG and inactivates MGL <ref name="Bertrand" /> | With SAR629 interacting with the catalytic triad it inhibits the triad from breaking down 2-AG and inactivates MGL <ref name="Bertrand" /> | ||
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== Catalytic triad == | == Catalytic triad == | ||
MGL has a classic <scene name='58/580298/Catalytic_triad/1'>catalytic triad</scene> that contains Ser-His-Asp | MGL has a classic <scene name='58/580298/Catalytic_triad/1'>catalytic triad</scene> that contains Ser-His-Asp. The triad was found using site-directed mutagenesis of each individual residue and each of these amino acid residues are catalytically essential to MGL <ref name="Bertrand" />. The catalytic triad is located in the binding pocket buried at the bottom of it in the oxyanion hole connected by a water molecule. | ||
[[Image:Catalytic_triad_binding_pocket.png|300px]] | [[Image:Catalytic_triad_binding_pocket.png|300px|thumb|'''Figure 4:''' The binding pocket of MGL with the catalytic triad (shown in red) buried in it.]] | ||
This triad has a natural attraction to Endocannabinoids, specifically 2-arachidonylglycerol (2-AG) <ref name="Bertrand" />. | This triad has a natural attraction to Endocannabinoids, specifically 2-arachidonylglycerol (2-AG) <ref name="Bertrand" />. | ||
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===Overall Reaction=== | ===Overall Reaction=== | ||
[[Image:Reaction.PNG|350px|thumb| | [[Image:Reaction.PNG|350px|thumb|'''Figure 5:''' The breakdown of 2-AG into arachidonic acid +glyverol by MGL.]] | ||
In this reaction 2-AG binds to the catalytic triad in the oxyanion hole in the active site. In the [[:Category:Oxyanion hole| oxyanion holes]] the substrate is stabilized by two nitrogen atoms from the catalytic Hisdidine and Asperatate during the transition step of the catalytic reaction. The catalytuc triad activates the nucleophilic serine and cleaves the ester bond of 2-AG that is being stabilized by its carbonyl group that is attached to the oxyanion hole. The glycerol molecule is released and it might diffuse to the narrow "exit hole", while the arachidonic acid would diffuse back to the top of the tunnel and leave the protein <ref name="Bertrand" />. | In this reaction 2-AG binds to the catalytic triad in the oxyanion hole in the active site. In the [[:Category:Oxyanion hole| oxyanion holes]] the substrate is stabilized by two nitrogen atoms from the catalytic Hisdidine and Asperatate during the transition step of the catalytic reaction. The catalytuc triad activates the nucleophilic serine and cleaves the ester bond of 2-AG that is being stabilized by its carbonyl group that is attached to the oxyanion hole. The glycerol molecule is released and it might diffuse to the narrow "exit hole", while the arachidonic acid would diffuse back to the top of the tunnel and leave the protein <ref name="Bertrand" />. | ||
==Ligand Binding Site== | ==Ligand Binding Site== | ||
[[Image:Overall_ligand.png|left|200px|thumb|Ligand within the Overall Structure of MGL]] | [[Image:Overall_ligand.png|left|200px|thumb|'''Figure 6:''' Ligand within the Overall Structure of MGL]] | ||
The <scene name='58/580298/Ligand/1'>ligand binding pocket</scene> of MGL has a large hydrophobic region with a polar bottom. The entrance of the binding pocket for MGL contains a lid, which is very flexible. The binding pocket or tunnel within MGL matches with the overall structure of 2-AG, with 2-AG's polar head being cleaved by the catalytic triad. Bertrand found that in MGL the binding pocket is not adjusted to the ligand's shape. However, the main movements of MGL associated with ligand binding involved the lid region. When 2-AG and its isomer 1(3)-AG bind to MGL, the hydrophobic chain is first aligned with the left part of the binding pocket. The carbonyl is then hydrogen bonded to <scene name='58/580298/Ala61/1'>Ala61</scene>. The polar head group of the ligand is then fixed by three hydrogen bonds. As a result, future research is looking into the large lipophilic portion of the binding pocket for designing selective inhibitors <ref name="Bertrand" />. | The <scene name='58/580298/Ligand/1'>ligand binding pocket</scene> of MGL has a large hydrophobic region with a polar bottom. The entrance of the binding pocket for MGL contains a lid, which is very flexible. The binding pocket or tunnel within MGL matches with the overall structure of 2-AG, with 2-AG's polar head being cleaved by the catalytic triad. Bertrand found that in MGL the binding pocket is not adjusted to the ligand's shape. However, the main movements of MGL associated with ligand binding involved the lid region. When 2-AG and its isomer 1(3)-AG bind to MGL, the hydrophobic chain is first aligned with the left part of the binding pocket. The carbonyl is then hydrogen bonded to <scene name='58/580298/Ala61/1'>Ala61</scene>. The polar head group of the ligand is then fixed by three hydrogen bonds. As a result, future research is looking into the large lipophilic portion of the binding pocket for designing selective inhibitors <ref name="Bertrand" />. | ||