Lipase

Introduction Lipase is a hydrolase that catalyzes the breakdown of lipids by hydrolyzing the esters of fatty acids. Lipases are important in digestion, promoting absorption of fats in the intestines. Lipase is primarily found in and secreted by the pancreas but is also found in the saliva and the stomach. Pancreatic lipase (PDB ID: 1HPL) which is pictured below is a carboxylic ester hydrolase. It is also commonly called pancreatic triacylglycerol lipase and its enzyme class number is E.C. 3.1.1.3 . The reaction catalyzed by this enzyme is shown below. Further breakdown ultimately results in 2-monoacylglycerols and free fatty acids . Pancreatic liapase is a 50 kDa protein, consisting of two identical, 449 residue chains . The determination of the structure and function of lipase was a gradual process. Lipase activity was first demonstrated in the pancreas by Claude Bernard in 1846. It wasn't until 1955 that Mattson and Beck demonstrated a high-specificity of pancreatic lipase for triglyceride primary esters . In recent years, determination of the crystal structure of pancreatic lipase has become the focus and many scientists have worked to further this. See also Molecular Playground/Pancreatic Lipase. Structure The s of lipase (in one subunit) include 102 residues which create 13 alpha helices, shown in red, and 139 residues involved in beta sheets totaling 28 strands, shown in gold. The alpha helices account for 22% of the protein, while the beta sheets comprise 30%. Each chain contains two well defined . The N terminal domain, shown in blue, is characterized by an alpha/beta hydrolase fold. While the C terminal domain, shown in green, contains a beta sheet sandwich which interacts with colipase . Each monomer and dimer structure of lipase is held together by disulfide bonds, hydrogen bonds, and electrostatic interactions (salt bridges). Lipase has 12 total between cysteine residues. are formed between the positively charge nitrogens (blue) in Arg and Lys, and negative oxygens (red) in Asp and Glu residues. also stabilize the enzyme and . Lipase has a distinct distribution of hydrophobic and hydrophilic residues. Hydrophobic collapse contributes to much of the secondary and tertiary structures, as the , shown in grey, point towards the interior of the protein. Conversely, the , in pink, point outwards . In addition, lipase has two , one buried in each monomer subunit. The image shows the green calcium ion in subunit A, coordinated by Glu187, Arg190, Asp192, and Asp195. The Ca(+2) charge is stabilized by negatively charged glutamate and aspartate residues, and the oxygen atoms from two water molecules (pink). The calcium ion is essential to protein folding and enzyme activity . The N-terminal domain also contains a that blocks solvent from entering the active site. Lipase Catalytic Mechanism Although a diverse array of lipase enzymes are found in nature, occupying diverse protein scaffolds, most are built upon an alpha/beta hydrolase fold and possess a chymotrypsin-like comprised of an acidic residue, a histidine, and a serine nucleophile. In the case of the images above of a horse pancreatic lipase, the catalytic triad is comprised of This catalytic triad functions like most found in nature, first with the Aspartic acid forming a hydrogen bond with His 263, increasing the pKa of the histidine imidazole nitrogen. This allows the histidine to act as a powerful general base and deprotonate the serine. The deprotonated serine then can serve as a nucleophile and attack the ester carbonyl of one of the fatty acids on the 1 or 3 carbons of the glycerol backbone of the lipid substrate. Upon attacking the lipid, a negatively charged tetrahedral intermediate is formed (Reaction 1). It is stabilized in the oxyanion hole by two residues: . The carbonyl reforms with the glycerol backbone segment acting as the leaving group (Reaction 2). A water molecule then donates a proton to the histidine, creating a reactive hydroxyl anion, which can attack the carbonyl carbon of the lipid, forming another negatively charged tetrahedral intermediate which is stabilized in the oxyanion hole (Reaction 3). Upon reformation of the carbonyl, the catalytic serine is released and monoglyceride and fatty acid monomers diffuse away (Reaction 4). Reaction 1: File:M0218.stg01.gif Reaction 2: File:M0218.stg02.gif Reaction 3: File:M0218.stg03.gif Reaction 4: File:M0218.stg04.gif Inhibition of Pancreatic Lipase In this structure, only one of the two identical chains is shown for lipase and colipase to better visualize the interaction of substrates and ligands with the protein. , a C11 alkyl phosphonate, is a competitive inhibitor of pancreatic lipase which binds to the active site. It is highlighted in purple. There are also five B-octylglucoside molecules in association with lipase. They are shown in grey and red. MUP forms hydrogen bonds with : Ser 152 and His 263, which are part of the catalytic triad, and Phe 77 and Leu 153 which are the stabilizing residues located in the oxyanion hole . Protein - Substrate Interactions Lipase binds with numerous hydrophobic contacts. As is seen here the lipase interacts with the alkyl group of cholesteryl linoleate via a hydrophobic rift within the protein. This rift orients the molecule to optimize the lipolysis reaction. Shown in this scene is lipase from Candida rugosa in with two molecules of cholesteryl linoleate (grey). The active site residues including SER152, ASP176, and HIS263 are shown in red stick representation. Clinical Significance Pancreatic lipase is secreted into the duodenum through the duct system of the pancreas. In a healthy individual, it is at very low concentration in serum. Under extreme disruption of pancreatic function, such as pancreatitis or pancreatic cancer, the pancreas may begin to digest itself and release pancreatic enzymes including pancreatic lipase into serum. Measurement of serum concentration of pancreatic lipase can therefore aid in diagnosis of acute pancreatitis.

spinqualitylabelsall models Structure of HMG-CoA reductase (PDB entry 1hpl) Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

3D Structures of Lipase3D Structures of Lipase

Update November 2011

Eukaryote natives:Eukaryote natives:

1hpl – hLip – horse
1hlg – hLip – human - gastric
3jw8, 3hju – mono-glyceride hLip
1jmy – hBSSL
1akn – cBSSL – cattle
2bce - cBSSL (mutant)
1f6w - cBSSL – catalytic domain
3o0d – Lip – Yarrowia lipolytica
1gpl – Lip – Guinea pig

Prokaryote natives:Prokaryote natives:

3guu, 1lbs, 1lbt, 1tca, 1tcb, 1tcc – CaLipA – Candida antarctica
2veo – CaLipA – closed state
3icv – CaLipB (mutant)
1gz7, 1lpm, 1lps– CrLip 2 – Candida rugosa - closed state
1crl, 1trh – CrLip – open state
1llf – Lip – Candida cylindracea
3g7n – Lip - Penicillium expansum
1tia - Lip – Penicillium camemberti
2qua, 2qub – LipA – Serratia marcescens
2hih – Lip – Staphylococcus hyicus
2fx5 – Lip – Pseudomonas mendocina
1yzf – Lip – Enterococcus faecalis
1dt3, 1dt5, 1dte, 1du4, 1ein, 1tib – TlLip - Thermomyces lanuginose
1jfr – Lip – Streptomyces exfoliates
1oil – BcLip - Burkholderia cepacia
2lip – BcLip – open state
1cvl – Lip – Chromobacterium viscosum
1lgy – Lip II – Rhizopus niveus
1tic - Lip – Rhizopus oryzae
1thg – Lip – Geotrichum candidum
3tgl, 4tgl, 1tgl – RmLip– Rhyzomucor miehei
2zvd – PsLip - Pseudomonas sp. – open state
2z8x - PsLip – extracellular
2zj6, 2zj7 – PsLip (mutant)
2z8z – PsLip(mutant) – closed state
3lip, 3a6z - Lip - Pseudomonas cepacia – open state
1qge, 1tah – Lip – Pseudomonas glumae
2w22 – Lip – Geobacillus thermocatenulatus
1ji3, 1ku0 – Lip – Bacillus stearothermophilus
1ah7 - Lip – Bacillus cereus
2qxt, 2qxu, 1isp, 1i6w - BsLip – Bacillus subtilis
3d2a, 3d2b, 3d2c, 1t2n, 1t4m, 3qmm - BsLip (mutant)
2ory – Lip – Photobacterium lypoliticum
2z5g, 2dsn – Lip T1 – Geobacillus zalihae
3p94 – Lip – Parabacteroides distasonis
3ngm – Lip – Gibberella zeae

Lipase/colipase complexes. The colipase is a co-enzyme whose binding to lipase optimizes the enzymatic activityLipase/colipase complexes. The colipase is a co-enzyme whose binding to lipase optimizes the enzymatic activity

1n8s – hLip+colipase II
1eth, 1lpa - Lip+colipase II - pig

Hormone-sensitive-lipases (LIPE) hydrolyze the first fatty acid of the triacylglycerol substrateHormone-sensitive-lipases (LIPE) hydrolyze the first fatty acid of the triacylglycerol substrate

3k6k – EstE7(LIPE) – metagenome library
3fak, 3dnm – EstE5(LIPE) – metagenome library
1evq – AaEst2(LIPE) – Alicyclobacillus acidocaldarius
1u4n – AaEst2(LIPE) (mutant)

Putative lipases; Proteins with unknown function but structural similarity to lipase obtained in structural genomics projects.Putative lipases; Proteins with unknown function but structural similarity to lipase obtained in structural genomics projects.

2rau - Lip – Sulfolobus solfataricus
3bxp, 3d3n - Lip – Lactobacillus plantarum
3e0x - Lip – Clostridium acetobutylicum
1z8h – Lip – Nostoc sp. PCC 712
1vj3 - Lip – Nostoc sp.
3bzw – Lip - Bacteroides thetaiotaomicron
2pbl – Lip - Silicibacter

Lipase + inhibitorsLipase + inhibitors

3jwe, 3pe6 - mono-glyceride hLip + SAR629 – covalent inhibitor
3l1h – EstE5(LIPE)+FeCl3 – noninvasive inhibitor
3l1i, 3l1j - EstE5(LIPE)+CuSO4 – noninvasive inhibitor
3lij - EstE5(LIPE)+ZnSO4– noninvasive inhibitor
3h18, 3h17 - EstE5 (LIPE)+PMSF
3h19, 3h1b, 3h1a – EstE5 (LIPE)+methyl alcohol
3h1a – EstE5 SLIPE)+ethyl alcohol
3h19 – EstE5 SLIPE)+isopropyl alcohol
3g9t, 3g9u - EstE5 (HSLIPE)+p-nitrophenyl butyrate
3g9z - EstE5 (LIPE) +p-nitrophenyl caprylate
2nw6 – BcLip+ S inhibitor
4lip, 5lip, 1r4z, 1r50 – BcLip+ Rc-(Rp,Sp)-1,2-dioctylcarbamoyl-glycero-3-O-phosphonate
1r4z – BsLip+Rc-IPG-phosphonate
1r50 - BsLip+Sc-IPG-phosphonate
1k8q - Lip+phosphonate – dog
1ex9 – Lip+Rc-(Rp,Sp)-1,2-dioctylcarbamoyl-glycero-3-O-phosphonate – Pseudomonas aeruginosa
5tgl – RmLip+N-hexyl-phosphonate
1lpb – Lip (pig)+colipase+C11 alkyl phosphonate
3icw – CaLipB (mutant) +methyl hydrogen R hexylphosphonate
3a70 – PsLip+diethyl phosphate

Lipase conjugated with analogs to its reaction intermediatesLipase conjugated with analogs to its reaction intermediates

1lpn, 1lpo, 1lpp – CrLip+ sulfonates
3rar – CrLip+ phosphonate
1qz3 – EaEst2(mutant) (LIPE)+hexadecanesulfonate

Lipase showing bile-salt binding siteLipase showing bile-salt binding site

1aql – cBSSL+taurocholate

Lipase with substrate bound at active siteLipase with substrate bound at active site

2zyh – AfLip (mutant)+fatty acid – Archaeoglobus fulgidus
2zyi - AfLip+fatty acid+Ca
2zyr - AfLip+fatty acid+Mg
2zys - AfLip+fatty acid+Cl
1gt6 – TlLip+oleic acid - lipid ligand

Lipase conjugated to transition-state analogs showing the binding mode of the enzyme catalysisLipase conjugated to transition-state analogs showing the binding mode of the enzyme catalysis

1ys1 – BhLip+hexylphosphonic acid (R) 2-methyl-3-phenylpropyl ester
1ys2 – BhLip+hexylphosphonic acid (S) 2-methyl-3-phenylpropyl ester
1hqd – Lip+1-phenoxy-2-acrtoxy butane – Pseudomonas cepacia

Lipase+lipase chaperoneLipase+lipase chaperone

2es4 – Lip+lipase chaperone C-terminal - Burkholderia glumae