Electrostatic Potential (ESP) Measure polarization Electron Map density Electron distribution
Dipole Moment Measure bond length/angle
Measure bond strength
Organic software for 3D model
Click here download Rasmol
Click here download PyMol Click here download Jmol
Click here Chem EDDL
Click here chemical search. Click here CRC database
Modelling and 3D representation
Chemistry Database
Click here Spectra database(OhioState) Click here Spectra database (NIST)
Click here chem finder.
Spectroscopic Database
Click here down Swiss PDB
Modelling and 3D representation
Click here crystallography database.
✓ ✓
Click here NIST data
✓ Click here download Arguslab
Click here chem axon
Click here download Avagrado
Click here chem EdDL
Click here download chimera ✓
Measure polarization Electron Map density Electron distribution
Electrostatic Potential (ESP) Dipole Moment
Measure bond length/angle Measure bond strength
Organic software for 3D model
Click here download Rasmol
Click here download PyMol Click here download Jmol
Click here Chem EDDL
Click here chemical search.
Modelling and 3D representation
Quick Chemistry Database Check
Click here down Swiss PDB
Modelling and 3D representation
✓ ✓
Click here NIST data
✓ Click here download Arguslab
Click here chemaxon quick chem check
Click here download Avagrado
Click here chem EdDL
Click here for Visualization/3D sources
Click here download Marvin Sketch
Click here quick chemical check
Click here quick chemical check
Measure bond length/angle Measure number H2 bonds
Measure bond strength Protein 1, 2 , 3O structure
Presence of disulfide bond Presence alpha and beta pleated sheet
Protein Data Bank Protein database key in - PDB 4HHB
Click here Chimera tutorial
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2
Uses molecular modelling
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2
Chemical viewer 3D structure (Chimera)
Download PDB text file
File – fetch by ID- 4HHB
Select – residue – HEM Select – chain A – Action – Ribbon – Hide Select – chain B,C,D - Action – ribbon Hide Display only ligand Heme Tool- structural analysis - Distance Select 2 atom -by press control/shift/left click select 2 points Tool – structure analysis – create to get distance
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Check here 4HHB Chimera 1MBO Select Histidine that are close to ring Locate His F8 and E7 Make measurement
Click here download chimera
Tool – Sequence – choose sequence for 4 chains Identify amino acids of interest
4
Type PDB code – 4HHB Right click – select Hetero Select - HETATM – HEM 4 Heme is display from 4 chains
Measure bond length/angle Measure number H2 bonds
Measure bond strength Protein 1, 2 , 3O structure
Presence of disulfide bond Presence alpha and beta pleated sheet
Click here J mol protein video
Chemical viewer 3D structure (Jmol)
Uses molecular modelling
1
J mol executable file
Measure distance
Select measure – distance for porphyrin ring Measure ring size/distance Fe from plane Select protein – by residue – Histidine Measure and locate His F8 Measure and locate His E7
final heme – click here
J mol executable file
1
Type 4HHB into protein data bank Look for ligand Heme
Model kit to design molecule
Click here deoxyhemoglobin chimera
2 2
3
4
3
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final product All histidine shown
Get structure from PDB and MOL
Measure bond length/angle Measure number H2 bonds
Measure bond strength Protein 1, 2 , 3O structure
Presence of disulfide bond Presence alpha and beta pleated sheet
Organic software for 3D model (Pymol)
1 1
Click here - Protein Data Bank Protein database key in - PDB 4 letter code
3
Click here download PyMol
Click here Pymol video tutorial
Click file – open your download pdb file from Protein Data bank Get to command term – Type fetch 4HHB H - Hide – S - Show cartoon – C – Type by ss
Select 4 Hem – Look for 4 Hemes Select 4HHB – H – hide everything Select Heme – Show stick Look His – select and name His F8 and His E7
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Press S – sequence at bottom screen. Right click – zoom in Select HEM - hemoglobin
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Uses molecular modelling
2
Select heme – right click – action – around 5A Look for His F8 and E7 around heme Make measure for distance Double click to display name of atom
Measure bond length/angle Measure number H2 bonds
Measure bond strength Protein 1, 2 , 3O structure
Presence of disulfide bond Presence alpha and beta pleated sheet
Protein Data Bank Protein database key in - PDB 4HHB
Click here Swiss PDB tutorial
1
2
Uses molecular modelling
1
2
Chemical viewer 3D structure (Swiss PDB)
Download PDB text file
File – open 4HHB pdb downloaded from databank
Window – Control panel Remove – side chain Select – Group kind – HETATM Display – stereo view Show only selected 4 Heme
Click here down Swiss PDB
Select – Group kind – Histidine Select – Residue – close to 2A Locate Histidine and make measurement
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Check for heme and Histidine only from control panel
Select Histidine that are close to ring Locate His F8 and E7 and make measurement
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Heme
Hemoglobin - 4 chain - 4 heme - 4 Fe 2+
Fe 2+
Heme (porphyrin)
Hemoglobin - 2 alpha chain - 144 amino acid - 2 beta chain - 146 amino acid
Myoglobin - 1 chain – 1 heme – 1 Fe2+
- 154 amino acids
Hemoglobin Myoglobin
Fe 2+
Heme (porphyrin)
PDB code files
Oxyhemoglobin – 1GZX, 1JY7 Deoxyhemoglobin – 1A3N, 4HHB, 2HHB, 1HBB, 1G9V, 101J Myoglobin – 4MBN, 3RGK, 5MBN Fetal hemoglobin – 1FDM Sickle cell Hemoglobin - 2HBS, 1NEJ Cytochrome (bovine) - 2B4Z Cytochrome (horse) - 1HRC Cytochrome (yeast) - 1YCC Cytochrome (human) - 3NWV
PDB file type for data analysis
1
Analyze using Chimera/Pymol/Swiss PDB/Jmol
1
Chimera Swiss PDB
Jmol Pymol
Fe in cytochrome 1 polypeptide chain
Fe 2+
Cytochrome c
Possible Research Question
Measuring using 3D modelling
Data Collection using 3D modelling
Data Collection using Database
Click here Jmol Click here PyMol
- What is the distance bet Fe and His E7 and F8, and are they the same for diff
heme found in hemoglobin/myoglobin/Cytochrome - Is His E7/F8 orientation similar for Hemoglobin, Myoglobin, Cytochrome - Is there any differences bet distance/position/orientation of porphyrin ring for
Hemoglobin, Myoglobin, Cytochrome - How is Fe2+ located, along or out of plane for Hemo/Myoglobin/Cytochrome - Is distance bet Fe and ligand N of porphyrin the same for
Hemoglobin/Myoglobin/Cytochrome - Is there any variation in terms of Fe and His E7/F8 for
Hemoglobin/Myoglobin/Cytochrome - Why His E7 and F8 are located in such a way across many different species? Is
their orientation highly conserved and why? - Similarity among cytochromes found in diff species of organism
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Click here Ensembl
Structural similarity and diff bet Hemoglobin/Myoglobin and Cytochrome
Myoglobin Hemoglobin Cytochrome
Hemoglobin Chimera Pymol Jmol Swiss PDB Mean
Orientation His/Fe Similar Similar Similar Similar Similar
Bond length N - Fe 2.12A 1.90A 2.02A 2.02A 2.01A
Bond length Fe – E7 5.93A 5.80A 5.45A 5.42A 5.55A
Bond length Fe – F8 2.25A 2.05A 2.10A 2.21A 2.13A
Chimera Swiss PDB
Data source
Myoglobin Chimera Pymol Jmol Swiss PDB Mean
Orientation His/Fe Similar Similar Similar Similar Similar
Bond length N - Fe 2.02A 2.11A 2.15A 2.32 2.14A
Bond length Fe – E7 5.80A 5.71A 5.56A 5.25A 5.25A
Bond length Fe – F8 2.15A 2.25A 2.11A 2.21A 2.21A
His E7
His F8
Fe
N
Possible Research Question Data Collection using 3D modelling
Data Collection using Database
Click here Jmol Click here PyMol
Click here NCBI Click here UCSC
Click here Ensembl
Structural similarity and differences bet Hemoglobin and Myoglobin
Myoglobin hemoglobin
vs
Chimera Swiss PDB
Evaluation and Limitation using 3D modelling
Must use a variety of sources/programme to verify/validate the validity and reliability of data collected Average is computed from diff software and checked with database to confirm. Check on methodological limitation using 3D model. (MUST perform 3D Optimization to most stable form structure. Critical and skeptical of result produced by computational chemistry. Major limitation of computation, they assume non-interacting molecule. (Ideal situation, ex molecule in vacuum or isolated state) Most appropriate molecule are those whose coordinates are not theoretical but derive from experimental structural determination (using X ray diffraction) Be careful of predicted arrangement from simulation /3D model Data sources are supported using diff method/3D model/database Certain database like NIST and CRC are more reliable source Check if there is a good agreement bet CRC, diff databases and 3D model prediction before making conclusion Computation programme is always based on approximation and we cannot conclusive prove anything Reflect of validity and reliability of data Is model a true representation of reality?
- What is the distance bet Fe and His E7 and F8, and are they the same for
diff heme found in hemoglobin/myoglobin - Is His E7/F8 orientation similar for Oxy, Deoxy and Myoglobin. - Is there any differences bet distance/position/orientation of porphyrin
ring for Hemoglobin and Myoglobin - How is Fe2+ located, along or out of plane for Hemo/Myoglobin - Is the distance bet Fe and ligand N of porphyrin the same for
Hemoglobin/Myoglobin - Is structure/size of porphyrin ring same for α and β chain - Is there any variation in terms of Fe and His E7/F8 for fetal hemoglobin
and sickle cell hemoglobin - Why His E7 and F8 are located in such a way across many different
species? Are their orientation highly conserved and why ?
- Porphyrin gp of heterocyclic made of 4 pyrrole subunit - Porphyrin macrocycle has 26 (delocalized) pi electron, obey Hückel rule - It is aromatic, 4n+2 π. (Highly conjugated system)
Heme
Porphyrin Heme = Fe + porphyrins ring
Heme
Heme A Heme B Heme C
Mitochondria - cytochrome c oxidase - electron transport
O2
Heme = Fe + porphyrin ring – carry O2
Fe2+ located
Most abundant Hemoglobin and Myoglobin
Mitochondria - cytochrome c - electron transport
Fetal Hemoglobin (2α22γ2)
Human Hemoglobin (2α2 2β2)
Sickle cell Hemoglobin (2α22βS
2) Myoglobin 1 α chain
Carbaminohemoglobin Carboxyhemoglobin Oxyhemoglobin
Heme
Heme A Heme B Heme C
Mitochondria - cytochrome c oxidase - electron transport
Most abundant Hemoglobin and Myoglobin
Mitochondria - cytochrome c - electron transport
Fetal Hemoglobin (2α22γ2)
Human Hemoglobin (2α2 2β2)
Sickle cell Hemo (2α22βS
2) Myoglobin 1 α chain
Cytochrome
Heme in cytochrome, highly conjugated ring sys surrounding Fe Cytochrome - REDOX rxn – mitochondria – ATP/energy production via elec transport chain Many type cytochromes – Cyto a, b, c1, a3 Cytochrome c, an ancient protein, developed early in the evolution of life. Essential protein for energy/ATP HIGHLY CONSERVED has changed little in millions of years. Many variation – but structure remain relatively unchanged
Fe in cytochrome Fe in cytochrome Cytochrome c – heme c
Hemoglobin A - 2 alpha and 2 beta chains Hemoglobin A2 - 2 alpha and 2 delta chains Hemoglobin F - 2 alpha and 2 gamma chains Heme (porphyrin) bind to Fe2+ using 4 nitrogen atom (histidine gp) Porphyrin
- as electron-pair donor - polydentate ligand Fe form 2 additional bonds, one on each side of the heme plane. These binding sites call fifth and sixth coordination sites. This hisitidine is referred as proximal Histidine F8 The sixth coordination site bind oxygen with His E7 nearby
Deoxyhemoglobin Fe2+ - out plane Can’t fit the ring
Heme
Hemoglobin - 4 chain - 4 heme (porphyrin) - 4 Fe 2+
Fe 2+
Heme (porphyrin)
Oxyhemoglobin Fe2+ - located in plane Fit the ring
Deoxyhemoglobin
Fe2+ out plane Can’t fit the ring Out by 0.06nm
Fe2+ in plane Fit the ring
Human hemoglobin - 2 alpha chain - 144 amino acid - 2 beta chain - 146 amino acid
Fe bind to six ligand. 4 with N atom of porphyrin Fifth ligand is donated by His F8 O2 add to Fe as sixth ligand O2 tilt relative to perpendicular of heme plane
His F8 His F8 His F8
His E7 His E7
His E7
Oxyhemoglobin
Heme
Hemoglobin - 4 chain - 4 heme (porphyrin) - 4 Fe 2+
Fe 2+
Heme (porphyrin)
Hemoglobin - 2 alpha chain - 144 amino acid - 2 beta chain - 146 amino acid
Fe bind to six ligand. 4 with N atom of porphyrin Fifth ligand is donated by His F8 O2 add to Fe as sixth ligand O2 tilt relative to perpendicular of heme plane
His E7 locate over Fe, force CO to bind to Fe at an angle. This steric hinderance reduce afinity of CO in hemoglobin. O2 bind to Fe at an angle, its binding not affected by presence of His E7.
His (E7)
His (F8)
vs
Myoglobin - 1 chain – 1 heme (porphyrin) - 1 Fe2+
- 154 amino acids
Hemoglobin Myoglobin
Fe 2+
Heme (porphyrin)
His (F8)
His (E7)