Human Gastric Lipase 1HLG

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Human Gastric Lipase (1HLG)

• Presented by Melanie Thomas, Ankur Patel, Brett
  Williams
• Biocomputing 3100

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I. Background Information

• Melanie Thomas

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Gastric Lipase

• Function Enzyme aiding in fat digestion
• Location of action Secreted by the stomach

www.bupa.co.uk/.../html/organ/stomach.html
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How Gastric Lipase Works

• Gastric lipase catalyzes the hydrolysis of stored
  triacylglycerols
• Substrate triacylglycerol
• Products long chain fatty acid and glycerol
• No cofactors

http//www.rpi.edu/dept/bcbp/molbiochem/MBWeb/mb2/
part1/lipoprot.htm
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How Gastric Lipase Works

• Hydrolyze substrate
• Present in intersurface between water and lipids
• Conformational Change
• Enzyme binds to intersurface ( between water and
  lipids)
• Activation occurs
• Active site of lipase is exposed
• Lipase binds to intersurface
• Helix moves
• Substrate binds at active site

http//www.albany-clinic.co.uk/weight-loss/index.p
hp?sessionb205u5sg
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Sources

• Organism
• Humans
• 1st mammalian lipase structure to be described
• Hydrolyzes 10-30 of fats in adults
• Most fats digested by pancreatic lipase
• Majority of mammals
• Rabbits
• Dogs
• Other Related Mammalian Lipases
• Lingual
• Rats

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Humans

• Premature infants
• 95 of dietary fat in human milk and infant
  formula is triglyceride
• Gastric lipase is main digestive enzyme
• 25-60 of total lipid digestion
• Pancreatic lipase is not fully developed
• Researching ways to modify enzyme to benefit
  babies born prematurely.

http//www.drexel.edu/univrel/digest/archive/02200
6/baby.jpg
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Humans

• Obesity in America
• Health problems
• Hypertension
• Diabetes
• High Cholesterol Levels
• Fats are most difficult part of diet to digest
• Anti-obesity drugs
• Suppress appetite

Microsoft Office Clip Art
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Diet Pills

• Aim to inhibit the action of digestive lipases
• Reducing fat absorption
• Blocks lipases active site
• Inhibiting triglyceride digestion

Microsoft Office Clip Art
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Diet Pills

• Orlistat
• Specifically targeted to inhibit gastric lipase
  and other digestive lipases
• Made from lipstatin
• Covalent inhibitor of digestive enzymes
• Much research many clinical trials
• Must be taken with a meal
• Inhibited 46.6 91.4 of activity of gastric
  lipase
• Reduced 11-33 of fat absorption by the stomach

Microsoft Office Clip Art
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Disease

• Deficiency of Lipase
• Health problems
• Gall Stones
• Heart Problems
• Minimum nutrient absorption
• Wolman Disease
• Cholesteryl Ester Storage Disease

Microsoft Office Clip Art
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Disease
Microsoft Office Clip Art

• Caused by storage of triglycerides in body
• Not being absorbed or removed
• Excess fats in feces
• Lipids build up in the cells, blood, and lymph
  and causes damage
• Many digestive system problems

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Special Applications

• Vitamins
• Patients with lipase deficiency
• Some Medical Conditions cause an absence of
  lipase
• Cystic Fibrosis Patients
• Post Surgery Patients
• Some Cancer Patients
• Supplements
• Corn has been engineered to produce gastric
  lipase
• Still a work in progress
• Under clinical trials

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II. Structure of Human Gastric Lipase

• Ankur Patel

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Crystal Structure Of Human Gastric Lipase

• PDB number 1HLG (Human Gastric Lipase)
• Method used to determine structure X-Ray
  diffraction

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General Structure Information

• Number of chains 2 (chain A and chain B)
• Both are identical
• Amino acid chains
• Each chain composed of 368 amino acids (excluding
  gaps)
• 379 amino acids with gaps included
• Gap 1 from amino acid 1 8 (8 residues)
• Gap 2 from amino acid 54 - 56 (3 residues)
• Total of 46 helices
• Total of 19 strands
• 2 total disulfide bridges

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Structure Showing Gaps 1 and 2

• Shows only chain A

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Helices

• CHOFAS

• Predicts 12 helices in chain A (24 total)

• GOR4

• Predicts 9 helices in chain A (18 total)

• PELE (JOI)

• Predicts 10 helices in chain A (20 total)

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Helices

• Cn3D

• Shows 11 helices in chain A (22 total)

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Helices

• Protein Explorer

• Shows 46 helices total (23 in each chain).
• Helices shown in pink

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Compiling information from Chofas, GOR4, PELE,
Cn3D and Protein explorer

• Notice that each method shows a different number
  of helices
• Chofas, GOR4 and Pele only PREDICT helices
• The prediction algorithms show fewer helices than
  Protein explorer

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Strands

• CHOFAS

• Predicts 17 strands in chain A (total of 34)

• GOR4
• Predicts 20 strands in chain A (total of 40)

• PELE (JOI)

• Predicts 19 strands in chain A (total of 38)

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Strands

• Cn3D

• Shows 8 strands per chain (16 total).
• 1 is anti-parallel, the remaining are parallel

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Strands

• Protein Explorer

• Shows a total of 16 strands (8 in each chain)
• Strands shown in yellow

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Compiling information from Chofas, Pele, GOR4,
Cn3d and Protein Explorer

• Biology workbench algorithms predict a high
  number of strands in each chain.
• The results from protein explorer and Cn3d show
  less strands

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Disulfide Bridges

• 2 total disulfide bridges
• Each is an intra-chain disulfide bridge
• Disulfide bridge between cysteine 227 and
  cysteine 236

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Ligand Animation
10 Ligands seen in the structure None are
directly associated
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Domains

• 2 Domains present
• Globular/core domain (colored purple)
• belongs to the alpha/beta hydrolase-fold family
• Between residues 9 189
• Between residues 307 379
• Cap domain (colored blue)
• Between residues 190 - 286

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• Alpha/beta hydrolase fold family
• All have a classical catalytic triad
• (Ser-153, His-353, Asp-324)
• Posses an oxayanion hole (stabilizes the
  oxayanion hole transition state).
• Hydrogen bonds between 2 main chain groups
• The cap domain has a lid.
• Covers catalytic SER-153
• SER-153 is therefore not freely accessible to
  substrates
• Lid must be displaced in order for substrate to
  bind to catalytic SER-153

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PROSEARCH

• Motif of interest Lipase_SER
• Confirms presence of catalytic triad
• Typical catalytictriad (Ser-153, His-353,
  Asp-324).
• The catalytic serine is deeply buried under a
  segment consisting of 29 residues the Lid.

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Lid of the Cap domain
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Active Sites

• One active site per chain in the core domain
• Residues SER 153 ASP 324 HIS 353.

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Active Site

• Note how lid blocks substrate from entering
  active site and associating with SER 153

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Dog Gastric Lipase (1K8Q)

• Structure has been crystallized with a phosponate
  inhibitor.
• C11 - Undecyl-Phosphinic Acid Butyl Ester
• Makes crevice
• BOG - B-Octylglucoside
• Fills up remainder of crevice

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Interaction of Inhibitor and Catalytic Serine
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Human (1HLG) vs. Dog Gastric Lipase (1K8Q)
Open conformation (inhibitor not shown)
Closed conformation
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TMAP, GREASE, TMHMM

• Reveals no transmembrane segment

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III. Phylogeny / Homology

• Brett Williams

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BLASTP
Canis lupus familiaris
www.webkinzinsider.com
Canis lupus
http//en.wikipedia.org/wiki/Gray_wolf
Bos taurus
Homo sapien
www.aspkin.com/.../2007/07/grazing-cow-1b.jpg
www.psxextreme.com/.../Christina_Aguilera.jpg
Mus musculus
http//en.wikipedia.org/wiki/Mus_musculus
Rattus norvegicus
www.orlandorats.com/images/bioNorway.jpg
Danio rerio
http//en.wikipedia.org/wiki/Zebrafish
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Unrooted Phylogenetic Tree
97
100
100
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Newick Format
((((Dog_Gastric_Lipase,Gray_Wolf)100,1HLG-Human_
Gastric_Lipase)97,Cow)82,(House_Mouse,Brown_Ra
t)100,Zebrafish)
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CLUSTALDIST Used to Create Scale Bar Smallest
Distance 0.005
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PROTPARS Unrooted Parsimonious Tree
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Human Gastric Lipase (1HLG) vs Dog Gastric
Lipase (1K8Q)
Missing residues Open/Closed Conformations
http//www.ncbi.nlm.nih.gov/Structure/vast/vastsrv
.cgi?sdid39042
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Polyhydroxyalkanoates (PHA)

• Analyzing the similarities b/w Human Gastric
  Lipase Type II Pseudomonas sp. USM 455 PHA
  synthase 1
• Catalyzed by PHA synthase
• Biodegradable polyesters made from microorganisms

www.biotech-weblog.com/50226711/biodegradable...
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Predicted PhaC1P.spUSM 4-55
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MSA including Pha C1 Synthase
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References

• Carriere,F., Renou, C., Ransac, S. (2001).
  Inhibition of gastrointestinal lipolysis by
  Orlistat during digestion of test meals in
  healthy volunteers Electronic version. AJP
  Gastrointestinal and Liver Physiology, 281,
  16-28.
• Pafumi, Y., Lairon, D., Lechenedela Porte, P.
  (2001). Mechanisms of Inhibition of
  Triacylglycerol Hydrolysis by Human Gastric
  Lipase Electronic Version. The Journal of
  Biological Chemistry, 277, 28070-28079.
• Roussel, A., Canaan, S., and Egloff, M. (1999).
  Crystal Structure of Human Gastric Lipase and
  Model of Lysosomal Acid Lipase, Two Lipolytic
  Enzymes of Medical Interest Electronic
  Version. The Journal of Biological Chemistry,
  274,16995- 17002.
• Wahab HA, Bahiyah N, Khairudin A, Samian MR,
  Najimudin N Sequence analysis and structure
  prediction of type II Pseudomonas sp. USM 4-55
  PHA synthase and an insight into its catalytic
  mechanism. BMC Structural Biology 2006, 623.
• http//www.ncbi.nlm.nih.gov/Structure/vast/vastsrv
  .cgi?sdid39042
• http//www.gate2biotech.com/polyhydroxyalkanoates-
  the-biodegradable-plastics/