Sandbox 51: Difference between revisions
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<applet load='1hpl' size=' | <applet load='1hpl' size='400' frame='true' align='right' caption='Lipase' /> | ||
==Introduction== | ==Introduction== | ||
Pancreatic lipase 1hpl (EC 3.1.1.3) is a an enzyme involved with the digestion and absorption of triacylglycerols (fats) in the intestine. It is secreted by the pancreas into the duodenum where it participates in the initial stages of breaking down fats into glyercol and three fatty acids. Lipase is a serine protease and a polar molecule. The mechanism of action requires Calcium ligands for stability and the coenzyme, colipase, for activation of the lipase mechanism. The hydrolysis products diffuse across enterocyte membrane and are absorbed into the blood <ref>Voet, D.,etc. "Fundamentals of Biochemistry: Life at the Molecular Level" John Wiley and Sons, Inc: New Jersey, 2008.</ref>. Due to that role lipase has in digestion of fat, and the growing problem of obesity, there has been increased effort to develop lipase inhibitors for weight loss supplements. | Pancreatic lipase 1hpl (EC 3.1.1.3) is a an enzyme involved with the digestion and absorption of triacylglycerols (fats) in the intestine. It is secreted by the pancreas into the duodenum where it participates in the initial stages of breaking down fats into glyercol and three fatty acids. Lipase is a serine protease and a polar molecule. The mechanism of action requires Calcium ligands for stability and the coenzyme, colipase, for activation of the lipase mechanism. The hydrolysis products diffuse across enterocyte membrane and are absorbed into the blood <ref>Voet, D.,etc. "Fundamentals of Biochemistry: Life at the Molecular Level" John Wiley and Sons, Inc: New Jersey, 2008.</ref>. Due to that role lipase has in digestion of fat, and the growing problem of obesity, there has been increased effort to develop lipase inhibitors for weight loss supplements. | ||
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==Mechanism== | ==Mechanism== | ||
In addition to the effects of bile salts, Lipase is activated by the coenzyme colipase, which binds to the C-terminal non-catalytic domain of lipase. Upon binding, active lipase is stabilized for the hydrophobic interaction with the triacylglycerides <ref>Fundamentals of Biochemistry...</ref>. Colipase must be present for activation of lipase and acts as a bridge between lipase and the lipid <ref>Crandall,W., Lowe, M. "Colipase Residues Glu64 and Arg65 Are Essential for Normal Lipase-mediated Fat Digestion in the Presence of Bile Salt Micelles" Journal of Biological Chemistry, 2001, (276) 12505-12512</ref>. Colipase is also secreted in the pancreas, but in its inactive form, which must be activated by trypsin before interacting with lipase. Colipase is a small protein cofactor with 5 conserved disulfide bonds <ref>"Colipase". Wikipedia: The Free Encyclopedia. 5 July 2011 [http://en.wikipedia.org/wiki/Colipase]</ref>, and 2 surfaces- a hydrophilic surface (site of lipase C-terminal interaction) and a hydrophobic surface (contains multiple hydrophobic loops to bridge the lipid)<ref>"Colipase Residues..."</ref>. Colipase and lipase <scene name='Sandbox_51/Contacts/3'>contacts</scene> are opposite of the active site on the C-terminal. The enzymes are bound by polar interactions such as <scene name='Sandbox_51/Salt_bridges/2'>salt bridges</scene> and van der waals forces <ref>van Tilbeurgh H, etc."Structure of the pancreatic lipase-procolipase complex", 1992 Sep 10;359(6391):159-62. PMID:1522902.[http://www.proteopedia.org/wiki/index.php/1n8s]</ref>. Specifically, interactions at the <scene name='Sandbox_51/Colipase_lipase_interface/1'>lipase-colipase interface</scene> include bonding between residues Glu15 and Arg38 with the lipase polar lid; and Arg44, Glu45, Glu64, Arg65, and Asn89 residues with main chain, C-terminal lipase residues <ref>"Colipase Residues..."</ref>. | In addition to the effects of bile salts, Lipase is activated by the coenzyme colipase, which binds to the C-terminal non-catalytic domain of lipase. Upon binding, active lipase is stabilized for the hydrophobic interaction with the triacylglycerides <ref>Fundamentals of Biochemistry...</ref>. Colipase must be present for activation of lipase and acts as a bridge between lipase and the lipid <ref>Crandall,W., Lowe, M. "Colipase Residues Glu64 and Arg65 Are Essential for Normal Lipase-mediated Fat Digestion in the Presence of Bile Salt Micelles" Journal of Biological Chemistry, 2001, (276) 12505-12512</ref>. Colipase is also secreted in the pancreas, but in its inactive form, which must be activated by trypsin before interacting with lipase. Colipase is a small protein cofactor with 5 conserved disulfide bonds <ref>"Colipase". Wikipedia: The Free Encyclopedia. 5 July 2011 [http://en.wikipedia.org/wiki/Colipase]</ref>, and 2 surfaces- a hydrophilic surface (site of lipase C-terminal interaction) and a hydrophobic surface (contains multiple hydrophobic loops to bridge the lipid)<ref>"Colipase Residues..."</ref>. Colipase and lipase <scene name='Sandbox_51/Contacts/3'>contacts</scene> are opposite of the active site on the C-terminal (contacts are regions of pink and yellow, with water molecules shown in darker blue). The enzymes are bound by polar interactions such as <scene name='Sandbox_51/Salt_bridges/2'>salt bridges</scene> and van der waals forces <ref>van Tilbeurgh H, etc."Structure of the pancreatic lipase-procolipase complex", 1992 Sep 10;359(6391):159-62. PMID:1522902.[http://www.proteopedia.org/wiki/index.php/1n8s]</ref>. Specifically, interactions at the <scene name='Sandbox_51/Colipase_lipase_interface/1'>lipase-colipase interface</scene> include bonding between residues Glu15 and Arg38 with the lipase polar lid; and Arg44, Glu45, Glu64, Arg65, and Asn89 residues with main chain, C-terminal lipase residues <ref>"Colipase Residues..."</ref>. | ||
Lipase activation at the lipid-water interface of triacylglycerides, in the presence of colipase and bile salts, is known as interfacial activation. For the hydroloysis reaction to take place, colipase anchors lipase to the lipid-water membrane of the micelle and a surface change occurs on lipase. Colipase's 4 hydrophobic loops interact with the hydrophobic atmosphere of the triacylglyceride initiating the lipase active site binding to the lipid, and lid opening to reveal a more hydrophobic environment for the triacylglycerol. Once colipase is bound, lipase initiates a serine-like hydrolysis involving the His-Asp-Ser active site residues in the catalytic triad <ref>Fundamentals of Biochemistry...</ref>. In the reaction, serine attacks the ester, forming an acyl-enzyme intermediate. The His and Asp residues help to stabilize the oxyanion intermediate through hydrogen bonding. Water enters the active site and reacts to release lipase and free the fatty acid. This acylation and deacylation reaction is usually completely reversible. | Lipase activation at the lipid-water interface of triacylglycerides, in the presence of colipase and bile salts, is known as interfacial activation. For the hydroloysis reaction to take place, colipase anchors lipase to the lipid-water membrane of the micelle and a surface change occurs on lipase. Colipase's 4 hydrophobic loops interact with the hydrophobic atmosphere of the triacylglyceride initiating the lipase active site binding to the lipid, and lid opening to reveal a more hydrophobic environment for the triacylglycerol. Once colipase is bound, lipase initiates a serine-like hydrolysis involving the His-Asp-Ser active site residues in the catalytic triad <ref>Fundamentals of Biochemistry...</ref>. In the reaction, serine attacks the ester, forming an acyl-enzyme intermediate. The His and Asp residues help to stabilize the oxyanion intermediate through hydrogen bonding. Water enters the active site and reacts to release lipase and free the fatty acid. This acylation and deacylation reaction is usually completely reversible. | ||
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[[Image:F6.medium.gif]] <ref> Reetz, Manfield F. Controlling the enantioselectivity of enzymes by directed evolution: Practical and theoretical ramifications. PNAS: 12 April 2004 [http://www.pnas.org/content/101/16/5716.full]</ref>. | [[Image:F6.medium.gif]] <ref> Reetz, Manfield F. Controlling the enantioselectivity of enzymes by directed evolution: Practical and theoretical ramifications. PNAS: 12 April 2004 [http://www.pnas.org/content/101/16/5716.full]</ref>. | ||
<applet load='1hpl' size=' | <applet load='1hpl' size='230' frame='true' align='left' caption='Lipase' /> | ||
==Inhibition== | ==Inhibition== | ||
The inhibition of pancreatic lipase has serious effects on storage and absorption of fats taken in by the body, and is therefore a potentially strong basis for pharmaceuticals to combat obesity. Because lipase is a part of the serine esterase family, it is inhibited in a similar manner. One such compound is a <scene name='Sandbox_51/Inhibitor/2'>C11 alkyl phosphonate</scene> inhibitor (shown interacting with the lipase-coplipase structure). The inhibitor acts by binding <scene name='Sandbox_51/Inhibitor_at_ser152/3'>Ser152</scene> in the active site inducing conformational changes in the beta-5 loop structures. The alkyl chain fits into the hydrophobic portion of the active lipase-colipase complex mimicking the fatty acid produced through hydrolysis in the normal enzymatic reaction <ref>Egloff, M.P., etc. "The 2.46 angstroms resolution structure of the pancreatic lipase colipase complex inhibited by a C11 alkyl phosphonate."(1995) J. Biochemistry 34: 2751-2762 [http://www.pdb.org/pdb/explore/explore.do?structureId=1lpb]</ref>. Van der waals forces between hydrophobic residues (blue) in the groove stabilize C11P binding, in addition to salt bridging and Hydrogen bonding forces with a cluster of hydrophilic residues (pick) around the Ser152 residue (purple) in the <scene name='Sandbox_51/Inhibitor_pocket/2'>inhibitor binding groove</scene>. <ref>Egloff, M., Marguet, F., Buono, G.,Verger,R.,Cambillau,C., Tilbeurgho,H. The 2.46 A Resolution Structure of the Pancreatic Lipase-Colipase Complex Inhibited by a C11 Alkyl Phosphonate? Biochemistry, 1995, 34, 275 1-2762. http://pubs.acs.org.ezproxy.messiah.edu/doi/pdf/10.1021/bi00009a003</ref>. Due to lipase's activity in the digestion and absorption of fat, there has been a growing market for lipase inhibitors for weight loss pharmaceuticals. The most popular is Orlistat (or Xenical®) which is a natural product from ''Streptomyces toxytricini'' and is the hydrogenation product of lipostation- an irreversible lipase inhibitor. This inhibitor also acts by binding Ser152, producing an ester | The inhibition of pancreatic lipase has serious effects on storage and absorption of fats taken in by the body, and is therefore a potentially strong basis for pharmaceuticals to combat obesity. Because lipase is a part of the serine esterase family, it is inhibited in a similar manner. One such compound is a <scene name='Sandbox_51/Inhibitor/2'>C11 alkyl phosphonate</scene> inhibitor (shown interacting with the lipase-coplipase structure). The inhibitor acts by binding <scene name='Sandbox_51/Inhibitor_at_ser152/3'>Ser152</scene> in the active site inducing conformational changes in the beta-5 loop structures. The alkyl chain fits into the hydrophobic portion of the active lipase-colipase complex mimicking the fatty acid produced through hydrolysis in the normal enzymatic reaction <ref>Egloff, M.P., etc. "The 2.46 angstroms resolution structure of the pancreatic lipase colipase complex inhibited by a C11 alkyl phosphonate."(1995) J. Biochemistry 34: 2751-2762 [http://www.pdb.org/pdb/explore/explore.do?structureId=1lpb]</ref>. Van der waals forces between hydrophobic residues (blue) in the groove stabilize C11P binding, in addition to salt bridging and Hydrogen bonding forces with a cluster of hydrophilic residues (pick) around the Ser152 residue (purple) in the <scene name='Sandbox_51/Inhibitor_pocket/2'>inhibitor binding groove</scene>. <ref>Egloff, M., Marguet, F., Buono, G.,Verger,R.,Cambillau,C., Tilbeurgho,H. The 2.46 A Resolution Structure of the Pancreatic Lipase-Colipase Complex Inhibited by a C11 Alkyl Phosphonate? Biochemistry, 1995, 34, 275 1-2762. http://pubs.acs.org.ezproxy.messiah.edu/doi/pdf/10.1021/bi00009a003</ref>. Due to lipase's activity in the digestion and absorption of fat, there has been a growing market for lipase inhibitors for weight loss pharmaceuticals. The most popular is Orlistat (or Xenical®) which is a natural product from ''Streptomyces toxytricini'' and is the hydrogenation product of lipostation- an irreversible lipase inhibitor. This inhibitor also acts by binding Ser152, producing an ester which hydrolyzes so slow that it is practically irreversible <ref>Kordik, C., Reitz, A. "Pharmacological Treatment of Obesity: Therapeutic Strategies" Journal of Medicinal Chemistry, 1999 (42).</ref>. | ||
==References== | ==References== | ||
<references /> | <references /> | ||
