1) General protein structure analysis 2) Problem 1: HIV-1 protease inhibitors 3) More examples are planned, ...To use the teaching module, proceed as follows:
Start WHAT IF.
Execute once the general command TEACH to copy all files from the `teach` directory (this is a subdirectory of the directory in which the WHAT IF executable resides) to your local directory. After that, type
FULLST Yto get out of WHAT IF again. (Dont worry about not understanding what you just did, it only needs to be done once, and it has nothing to do with science). Now you are ready to start running the teaching scripts.
Run the teaching scripts. Every script has a code name that is given in brackets at the end of the chapter heading (E.g. AA0001, BB0105, etc). So,
whatif @AA0001Runs the first script on SG machines. Under PCDOS you first start WHAT IF and use the script command like:
script AA0001
At the end of every script, WHAT IF terminates. So, for every script you have to start WHAT IF again.
Script names consist of 2 characters and two times two digits. These 3 groups generate three hierarchial layers in the scripts. The first two characters indicate:
AA General BB Problem 1 HIV-1 protease inhibitorsThe further division follows the lines of the table below:
AA GENERAL 00 Visualisation 01 Shows some display modes using Hypothase (all, C-alpha, H-bonds, Sec.Str., Acc.Surf.) 02 Shows some colouring schemes for the display of Hypothase residues 03 Emphasize that colouring schemes can identify differences in protein structure 04 Shows a Ramachandran plot of Hypothase 05 Shows contacts in Crambin by dashed lines 06 Shows contact plot of Crambin 07 Shows surface plots of HLA 08 Structural superposition (Hemoglobin A and B) 09 Shows some colouring schemes for the display of Hypothase residues (backbone, side-chains) 10 Shows surface plot of Hypothase(The AA 00 ** group holds the scripts used at the EMBL for a one day course for first year PhD students)
01 Primary structure 01 Primary structure of Hypothase 02 Classes of amino acids (example: Hypothase) 02 Secondary structure 01 Shows secondary structure (alpha helix) example 02 Shows secondary structure (anti-parallel beta sheet) example 03 Shows secondary structure (alpha helix) example including hydrogen bonding pattern 04 Shows secondary structure (anti-parallel beta sheet) example, including hydrogen bonding pattern 05 Shows secondary structure (parallel beta sheet) example 06 Shows secondary structure (parallel beta sheet) example including hydrogen bonding pattern 07 Ramachandran plot of Hypothase 03 Tertiary structure 01 Shows combination of secondary structures (TIM) including hydrogen bonding pattern and ribbons 02 Shows the structure of HIV-1 Protease sub-unit A including hydrogen bonding pattern and ribbons 03 Shows the structure of Hypothase including hydrogen bonding pattern and ribbons 04 Example of anti-parallel beta strands in the domains of Aspartate Transcarbamylase 05 Example of parallel beta strands in Flavodoxin 06 Example of (anti)parallel beta barrel in Plastocyanin 07 Example of the helix-loop-helix motif in Calmodulin 08 Example of the hairpin-beta motif in Erabutoxin 09 Example of the beta-alpha-beta-alpha motif in TIM 10 Example of the helix-bundle motif in Myohemerythrin 04 Quarternary structure 01 Shows the complete structure of Triose Phosphate Isomerase (TIM) including hydrogen bonding pattern and ribbons 02 Shows the complete structure of HIV-1 Protease including hydrogen bonding pattern and ribbons(The AA 00 01-04 group can be incorporated in any other set of scripts because they are very simple and very clear examples of elementary aspects of protein structure).
BB PROBLEM 1 HIV-1 PROTEASE INHIBITORS 00 Introduction 01 Structure of HIV-1 Protease 01 Primary structure (sub-unit A) 02 Tertiary structure (sub-unit A) (Backbone & C-alpha & H-bonds) 03 Tertiary structure (sub-unit A) (H-bonds & Ribbons) 04 Quaternary structure (sub-units A & B) (colours) 05 Quaternary structure (sub-units A & B) (Ribbons) 06 Shows tertiary structure of HIV-1 Protease with superposed sub-units A & B 07 Contacts between HIV-1 Protease sub-units 08 Plot of the contacts between the HIV-1 Protease sub-units 02 Homology of HIV-1 Protease with other structures 00 Introduction 01 Shows homology of HIV-1 Protease with Rouse Sarcoma virus 02 Shows homology of HIV-1 Protease with Penicillopepsin 03 Shows homology of HIV-1 Protease with Endothiapepsin 04 Shows homology of HIV-1 Protease with Rhizopuspepsin 05 Shows homology of Endothiapepsin with Penicillopepsin 06 Shows homology of Rhizopuspepsin with Endothiapepsin 07 Shows homology of HIV-1 Protease with Pepsin 03 Inhibition of HIV-1 Protease 01 Active site of HIV-1 Protease 02 Example of a HIV-1 Protease inhibitor (A74704) and its interaction with both sub-unit A & B 03 Contacts of inhibitor A-74704 with HIV-1 Protease active site(The BB 00 00-03 group is a two-three day course for third year undergraduate students at the University of Leiden. It is probably better to go through one or two days of AA scripts before starting the BB scripts. The BB scripts require sequence databases, Mosaic ot Netscape WWW browsing, etc., as support).
1) Visualisation 2) Primary structure 3) Secondary structure 4) Tertiary structure 5) Quarternary structure
MOL1 : the whole molecule, coloured by atom type, MOL2 : the hydrogen bonds present, MOL3 : the molecular surface, represented by dots, MOL4 : an alpha carbon trace, MOL5 : a backbone only trace, MOL6 : a ribbon representation.Questions:
1) Try to find the helix and the 3 strands.
2) Which hydrogen bond patterns govern these elements?
3) Why are these hydrogen bonds so much more irregular than expected?
RED : for acidic amino acids (glutamic and aspartic acid) BLUE : for basic amino acids (arginine, lysine, histidine) PURPLE : for polar residues (glutamine, asparagine) YELLOW : for sulphur containing amino acids (cysteine) GREEN : for hydrophobic residues (leucine, valine, tryptophan, etc.) GREENISH : for alcoholic residues (threonine, serine, tyrosine)The following graphical objects are available:
MOL1 : the whole molecule coloured by residue type MOL2 : as MOL1, but with the backbone reduced to a C-alpha trace MOL3 : as MOL1, but with the backbone reduced to a backbone trace MOL4 : a ribbon representationQuestions:
1) Are the polar and a-polar residues distributed randomly over the structure?
2) If not, derive a rule for this distribution.
3) Does this molecule have a 'hydrophobic core'?
MOL1 : the whole molecule coloured by residue type MOL2 : as MOL1, but with the backbone reduced to a C-alpha trace MOL3 : as MOL1, but with the backbone reduced to a backbone trace MOL4 : a ribbon representationQuestions:
1) Now that we know how residues are distributed over the protein, see how much of that rule holds in this molecule.
2) Any idea where the differences come from?
3) Does this molecule have a 'hydrophobic core'?
MOL1 : a Phi-Psi plot of Hypothase (click on a data point to identify the corresponding residue).Questions:
1) Try to find out which parts of the molecule end up where in this plot.
2) Three residues fall wide outside the 'boxed' areas. Why?
MOL1 : the whole molecule, coloured by atom type, MOL2 : dotted lines between contacting atoms, MOL3 : dots that indicate the Van der Waals surface.Questions:
1) Which residues are very important for the folding of this molecule?
2) Which residues are 'totally useless' for Crambin?
3) Why are those 'useless' residues nevertheless present?
MOL1 : a contact plot (click on the lower left corner of a square to identify the corresponding contacting residues).Questions:
1) Why is the diagonal of this plot sometimes wide, and sometimes narrow?
2) Can you find back the 'usefull' residues detected in the previous exercise?
MOL1 : the whole molecule, MOL2 : a very low resolution surface plot, MOL3 : a ribbon representation.Questions:
1) How many independend domains can you find in HLA?
2) Any idea where the oligo peptide probably sits in HLA?
1 - V L S P A D K T N V K A A W G K V G A H A G E Y G A E A L 30 V H L T P E E K S A V T A L W G K V - - N V D E V G G E A L 31 E R M F L S F P T T K T Y F P H F - D L S H - - - - - G S A 60 G R L L V V Y P W T Q R F F E S F G D L S T P D A V M G N P 61 Q V K G H G K K V A D A L T N A V A H V D D M P N A L S A L 90 K V K A H G K K V L G A F S D G L A H L D N L K G T F A T L 91 S D L H A H K L R V D P V N F K L L S H C L L V T L A A H L 120 S E L H C D K L H V D P E N F R L L G N V L V C V L A H H F 121 P A E F T P A V H A S L D K F L A S V S T V L T S K Y R * 150 G K E F T P P V Q A A Y Q K V V A G V A N A L A H K Y H *Around residue 50 we see a strong identical triplet: DLS. Based on the structures, Asp 49 of the A-chain should be aligned with Gly 48 from the B-chain.
Questions:
1) What is wrong in the above alignment around residue 50?
2) Try to improve the alignment near the n-termini.
PURPLE : for the backbone atoms. GREEN : for the side-chain atoms.The following graphical objects are available:
MOL1 : the whole molecule coloured by atom type. MOL2 : the whole molecule, the backbone and side-chain atoms coloured differently. MOL4 : Lines indicating atomic contacts MOL5 : A ribbon representationQuestions:
1) Try to analyse backbone-backbone, backbone-sidechain and sidechain-sidechain contacts.
2) What are the major differences between these three classes?
MOL1 : the whole molecule coloured by atom type. MOL2 : the surface map of the molecule.Questions:
1) Does this surface representation agree with the conclusions of AA0002?
2) Would you call the surface 'rather smooth', 'rather rippled', or would you give it another description.
3) Where sits the hypothase active site?
Questions:
1) Which is the smallest residue?
2) Which is the largest residue?
3) Which are the negatively charged residues?
4) Which are the positively charged residues?
5) Which are the alcoholic residues?
6) Which are the most hydrophobic residues?
7) Which are the most flexible residues?
8) Which are the most rigid residues?
acidic amino acids (Gln, Asp) RED basic amino acids (Arg, Lys, His) BLUE polar residues (Glu, Asn) PURPLE sulphur containing amino acids (Cys, Met) YELLOW small hydrophobic residues (Gly, Ala, Val, Pro) GREEN large hydrophobic residues (Leu, Trp, Phe, Ile) GREEN alcoholic residues (Thr, Ser, Tyr) GREENISHQuestions:
1) Label at least one residue from each class of residues.
MOL1 : An example of an alpha helix (poly-A). MOL2 : The backbone atoms of the alpha helix. MOL3 : The C-alpha atoms of the alpha helix.Questions:
1) How many residues are there in one helical turn?
MOL1 : An example of an anti-parallel beta sheet. MOL2 : The backbone atoms of the sheet, MOL3 : The C-alpha atoms of the sheet.Questions:
1) Why do we call this ANTIPARALLEL beta sheet?
MOL1 : An example of a parallel beta sheet. MOL2 : The backbone atoms of the sheet. MOL3 : The C-alpha atoms of the sheet.Questions:
1) Why do we call this PARALLEL beta sheet?
MOL1 : An example of an alpha helix (poly-A). MOL2 : The backbone atoms of the helix. MOL3 : The C-alpha atoms of the helix. MOL4 : The hydrogen bonds stabilizing the helix.Questions:
1) Is there any regularity in the hydrogen bonding pattern?
MOL1 : An example of an anti-parallel beta sheet. MOL2 : The backbone atoms of the sheet. MOL3 : The C-alpha atoms of the sheet. MOL4 : The hydrogen bonds stabilizing the sheet.Questions:
1) Is there any regularity in the hydrogen bonding pattern?
MOL1 : An example of a parallel beta sheet. MOL2 : The backbone atoms of the sheet. MOL3 : The C-alpha atoms of the sheet. MOL4 : The hydrogen bonds stabilizing the sheet.Questions:
1) Is there any regularity in the hydrogen bonding pattern?
2) What are the major differences in hydrogen bonding between parallel and antiparallel beta sheets?
Phi = N-Calpha rotation Psi = Calpha-C rotationA plot of Phi against Psi can be useful to get a quick impression of the secondary structure of a protein molecule. Because of steric hindrance not every combination of Phi & Psi torsion angles is possible. The allowed regions are indicated. The Phi/Psi plot of the protein Hypothase will be displayed:
MOL1 : a Phi-Psi plot of Hypothase (click on a cross to identify the corresponding residue).Questions:
1) Which areas in this plot correspond to alpha helix and beta strand?
MOL1 : The complete subunit A of TIM. MOL2 : The backbone atoms. MOL3 : The C-alpha atoms. MOL4 : The hydrogen bonds stabilizing helices, sheets and the subunit. MOL5 : A ribbon representation of the all secondary structure elements.Questions:
1) Follow the chain from N to C. Are there any regularities?
MOL1 : The complete sub-unit A of HIV-1 Protease. MOL2 : The backbone atoms. MOL3 : The C-alpha atoms. MOL4 : The dimer of HIV-1 Protease. MOL5 : A ribbon representation of the all secondary structure elements.Questions:
1) How many secondary structure elements in the A subunit make a contact with the B subunit?
2) Where do you think the amino acids will be located between which this protease cleaves.
MOL1 : The complete Hypothase. MOL2 : The backbone atoms. MOL3 : The C-alpha atoms. MOL4 : The hydrogen bonds stabilizing helices, sheets and the subunit. MOL5 : A ribbon representation of the all secondary structure elements.Questions:
1) Do you see the parallel and the antiparallel strands?
MOL1 : The complete structure of Aspartate Transcarbamylase. MOL2 : A ribbon representation of all the secondary structure elements.
MOL1 : The complete structure of Flavodoxin. MOL2 : A ribbon representation of all the secondary structure elements.Questions:
1) Follow the chain from N to C. How are the strands ordered?
MOL1 : The complete structure of Plastocyanin. MOL2 : A ribbon representation of all the secondary structure elements.
MOL1 : The complete structure of Calmodulin. MOL2 : A ribbon representation of all the secondary structure elements.
MOL1 : The complete structure of Erabutoxin. MOL2 : A ribbon representation of all the secondary structure elements.
MOL1 : The structure of TIM sub-unit A. MOL2 : A ribbon representation of all the secondary structure elements.
MOL1 : The structure of Myohemerythrin. MOL2 : A ribbon representation of all the secondary structure elements.
MOL1 : The complete structure of Triose Phosphate Isomerase (TIM). MOL2 : The backbone atoms. MOL3 : The C-alpha atoms. MOL4 : The hydrogen bonds stabilizing helices, sheets and the subunit. MOL5 : A ribbon representation of the all secondary structure elements.
MOL1 : The complete structure of HIV-1 Protease. MOL2 : The backbone atoms. MOL3 : The C-alpha atoms. MOL4 : The hydrogen bonds stabilizing helices, sheets and the subunit. MOL5 : A ribbon representation of the all secondary structure elements.
1) Structure of HIV-1 Protease 2) Homology of HIV-1 Protease with other structures 3) Inhibition of HIV-1 Protease
HIV-1 protease, with only 99 amino-acid resuidues, is the smallest of the retroviral proteases, and is much smaller than the microbial and mammalian aspartyl proteases, each of which contains approximately 325 residues.
1-50 PQITLWQRPL VTIKIGGQLK EALLDTGADD TVLEEMSLPG RWKPKMIGGI 51-99 GGFIKVRQYD QILIEICGHK AIGTVLVGPT PVNIIGRNLL TQIGCTLNFA representation of sub-unit A of HIV-1 Protease will be shown on the screen.
MOL1 : Sub-unit A of HIV-1 Protease. MOL2 : The backbone of sub-unit A. MOL3 : The C-alpha trace of sub-unit A. MOL4 : The H-bond pattern calculated for sub-unit A.
MOL1 : Sub-unit A of HIV-1 Protease. MOL2 : The backbone of sub-unit A. MOL3 : The C-alpha trace of sub-unit A. MOL4 : The H-bond pattern calculated for sub-unit A. MOL5 : A ribbon representation of sub-unit A.
MOL1 : HIV-1 Protease. MOL2 : The sub-units displayed in different colours. MOL3 : The backbone. MOL4 : The C-alpha trace. MOL5 : The calculated H-bond pattern.
MOL1 : HIV-1 Protease. MOL2 : The sub-units displayed in different colours. MOL3 : The backbone. MOL4 : The C-alpha trace. MOL5 : The calculated H-bond pattern. MOL6 : Ribbon representation of the molecule.
MOL1 : HIV-1 Protease, sub-unit A. MOL2 : HIV-1 Protease, sub-unit B. MOL3 : The superimposed sub-units A & B. MOL4 : Backbone sub-unit A. MOL5 : Backbone superposed sub-unit B. MOL6 : C-alpha trace sub-unit A. MOL7 : C-alpha trace superposed sub-unit B.
MOL1 : HIV-1 Protease, sub-unit A. MOL2 : HIV-1 Protease, sub-unit B. MOL3 : Contacts shown as dotted lines. MOL4 : HIV-1 Protease. MOL5 : Backbone sub-unit A. MOL6 : Backbone sub-unit B. MOL7 : C-alpha trace sub-unit A and B.
MOL1 : Plot of the contacts calculated between sub-unit A and B.
MOL1 : Sub-unit A of Rouse Sarcoma Virus Protease. MOL2 : Superposed sub-unit A of HIV-1 Protease. MOL3 : C-alpha trace of Rouse Sarcoma Virus Protease. MOL4 : C-alpha trace of superposed sub-unit A of HIV-1 Protease. MOL5 : The superposed sub-units.
This example shows the superposed HIV-1 sub-unit A upon Penicillopepsin.
MOL1 : Penicillopepsin. MOL2 : Superposed sub-unit A of HIV-1 Protease. MOL3 : C-alpha trace of Penicillopepsin. MOL4 : C-alpha trace of superposed sub-unit A of HIV-1 Protease. MOL5 : The superposed sub-unit on Penicillopepsin.
This example shows the superposed HIV-1 sub-unit A upon Endothiapepsin.
MOL1 : Endothiapepsin. MOL2 : Superposed sub-unit A of HIV-1 Protease. MOL3 : C-alpha trace of Endothiapepsin. MOL4 : C-alpha trace of superposed sub-unit A of HIV-1 Protease. MOL5 : The superposed sub-unit on Endothiapepsin.
This example shows the superposed HIV-1 sub-unit A upon Rhizopuspepsin.
MOL1 : Rhizopuspepsin. MOL2 : Superposed sub-unit A of HIV-1 Protease. MOL3 : C-alpha trace of Rhizopuspepsin. MOL4 : C-alpha trace of superposed sub-unit A of HIV-1 Protease. MOL5 : The superposed sub-unit on Rhizopuspepsin.
This example shows the superposed Penicillopepsin upon Endothiapepsin.
MOL1 : Endothiapepsin. MOL2 : Penicillopepsin. MOL3 : C-alpha trace of Endothiapepsin. MOL4 : C-alpha trace of superposed Penicillopepsin. MOL5 : The superposed Penicillopepsin on Endothiapepsin.
This example shows the superposed Rhizopuspepsin upon Endothiapepsin.
MOL1 : Endothiapepsin. MOL2 : Rhizopuspepsin. MOL3 : C-alpha trace of Endothiapepsin. MOL4 : C-alpha trace of superposed Rhizopuspepsin. MOL5 : The superposed Rhizopuspepsin on Endothiapepsin.
This example shows the superposed sub-unit A of HIV-1 Protease upon Pepsin.
By moving the molecule around, one is able to view through a hole in the structure which comprises the active site. As an example the surface area of a co-crystallized inhibitor is displayed to give an idea of the extent of the cavity.
MOL1 : HIV-1 Protease, sub-unit 1. MOL2 : HIV-1 Protease, sub-unit 2. MOL3 : Surface map of inhibitor A74704.
MOL1 : HIV-1 Protease, sub-unit A. MOL2 : HIV-1 Protease, sub-unit B. MOL3 : Inhibitor A74704. MOL4 : C-alpha trace sub-unit A. MOL5 : C-alpha trace sub-unit B.
MOL1 : HIV-1 Protease, sub-unit A. MOL2 : HIV-1 Protease, sub-unit B. MOL3 : Inhibitor A74704. MOL4 : C-alpha trace sub-unit A. MOL5 : C-alpha trace sub-unit B. MOL6 : Contacts for A-74704 with HIV-1 Protease.
Enzyme Inhibitor Sub-site Group Enzyme residues
S3 CBZ Gly27-A, Ala28-A, Asp29-A, Asp30-A, Gly48-A, Met46-A, Ile47-A S3' CBZ' Gly27-B, Ala28-B, Asp29-B, Asp30-B, Gly48-B, Arg8-A S2 VAL Ala28-A, Val32-A, Ile47-A, Gly48-A, Gly49-A, Ile50-A, Ile84-A S2' VAL' Ala28-B, Val32-B, Ile47-B, Gly48-A, Gly49-B, Ile50-B, Ile84-A, Asp29-B, Ile50-A S1 COR Leu23-A, Asp25-A, Gly27-A, Ala28-A, Gly49-A, Ile50-A, Val82-A, Ile84-A S1' COR' Leu23-B, Asp25-B, Gly27-B, Ala28-B, Gly49-B, Ile50-B, Val82-B, Ile84-B, Pro81-B, Arg8-A Central-OH Asp25-A, Gly27-A, Ala28-A Asp25-B, Gly27-B Buried H2O Gly49-A, Ile50-A, Gly49-B, Ile50-B VAL, VAL', COR (inhibitor groups)