Proteopedia - User contributions [en]

User:Nikhil Malvankar/IW2018

2019-09-09T22:53:54Z

Nikhil Malvankar:

<center>
How to visualize, understand and share/present 3D protein molecular structures 
by Nikhil Malvankar, 2019

 
Adapted from Eric Martz 
for
MB&B 420a/720a Macromolecular Structure Fall 2019
 
Yale University USA
</center>



==I. Getting Started==
#Use '''FIREFOX''' or Safari.(DO NOT USE Chrome because molecule rotation will be slow/jerky. Internet Explorer and Edge are even worse with this software.)
#Go to our syllabus.
#Now you can see this document in your browser. Go to '''[http://atlas.molviz.org Atlas.MolviZ.Org]'''.
#In the Atlas, choose any molecule deemed ''Straightforward'' and click on the link to '''FirstGlance'''. After a minute or so to load, you should see a rotating molecule.
#If you have any difficulty or the molecule does not appear, ask for help!

==II. Goals==

1. Review principles of protein 3D structure.

2. Choose an existing experimentally-determined 3D protein structure model to investigate.

3. Learn how ''FirstGlance in Jmol'' makes it easy to see structure-function relationships in the protein you chose.


4. Write a report including snapshots of your protein that illustrate your answers to the questions below. (Your report will be in the form of Slides emailed to TAs. You will not present your report in class.)

==III. Protein Structure and Structural Bioinformatics==
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[https://docs.google.com/presentation/d/1lS59hde5F6ur9VCBO1_eHwXbhQcUHByPPVi12w6PSRM/edit?usp=sharing '''SLIDESHOW''']
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:1. [[Amino acid]] '''sequence''' + protein chain '''conformation''' = protein '''function'''.
::A. [http://www.umass.edu/molvis/workshop/allstruc/whycare.htm Why do we care] about protein 3D structure?
::B. Conformation can be a '''stable fold''' or '''[[Intrinsically Disordered Protein|intrinsically unstructured]]'''. Both commonly exist in the same protein molecule.
::C. Conformation is specified by sequence.
:::*Folded domains fold spontaneously (Anfinson, 1960's<ref>For a brief overview of Anfinson's protein folding experiments in the 1960's, see the first paragraph at [[Intrinsically Disordered Protein]].</ref>), or with the help of [[chaperonins]].
:::*The '''denaturation''' (unfolding) of a folded protein domain destroys its function.

:2. Backbone Representation.

::A. [http://proteopedia.org/wiki/index.php/Backbone_representations Backbone Representations]
::B. [http://firstglance.jmol.org/fg.htm?mol=4d7b Small Protein in FirstGlance] (use the Views Tab: Vines, Cartoon)

:3. Structure Knowledge.

::A. Although sequence specifies fold, scientists '''cannot yet predict the fold from the sequence'''. Therefore, fold must be determined by empirical (experimental) methods. The most common methods for determining the 3D structure of a protein molecule are:
:::*[[X-ray crystallography]], 88%.
::::*Result is a single model representing the average of the molecules in the crystal.
::::*[[Resolution]] reflects the degree of order or disorder in the crystal.
::::*X-ray crystallography gives no models for [[Intrinsically Disordered Protein|intrinsically unstructured]] loops or molecules.
:::*[[Nuclear magnetic resonance]] (NMR) in aqueous solution, 9%.
::::*NMR is limited to small proteins (30 kD or smaller; median NMR in PDB is 10K; median X-ray is 50K).
::::*Result is an ensemble of models consistent with the data. Examples: [[2bbn]]
:::*High resolution cryo-electron microscopy, 0.8%.

::B. These methods are difficult and expensive. Less than 10% of proteins have known structure.
::C. All published, empirically determined 3D macromolecular structure models are available from the '''Protein Data Bank''' (PDB; [http://www.pdb.org pdb.org]; [[PDB|About the PDB]]).

::D. Each model has a unique, 4-character accession code called a '''PDB identification code''', for example
:::*[[1hho]]
:::*[[1d66]]
:::*[[2bbn]]
:::*[[9ins]]

::E. Crystallographers publish the [[asymmetric unit]] of the crystal. It may be identical with the [[biological unit]] (the functional form of the molecule), or it may be only part of the biological unit, or it may contain multiple copies of the biological unit. See [[Biological unit#Examples|examples]].
:::Interchain contacts that occur in the asymmetric unit, which are absent in the biological unit, are an artifact of crystallization, termed [[crystal contacts]].

==IV. Choose a Molecule to Explore==
*Choose a molecule to use for your report.
**Each student should choose a ''different'' molecule.
**Be sure to note the '''4-character PDB code''' of the molecule you choose. The PDB code makes it easy to retrieve the molecule and information about it.
**Report the PDB code you chose to the instructor to make sure it is not already taken.
**It must have protein.
**It will be more interesting if it contains some non-protein: ligand, or DNA or RNA.
**X-ray results should have resolution of 3 Å or better.
*Here are some ways to find a protein with known structure:
# '''Recommended:''' Atlas of Macromolecules ([http://atlas.molviz.org Atlas.MolviZ.Org]). Choose a "Straightforward" or "Challenging" (not "Enormous") molecule that has protein and ligand.
# [http://pdb101.rcsb.org/motm/motm-by-category Molecule of the Month] at the PDB. Look for PDB codes in the article, and use [http://firstglance.jmol.org FirstGlance] to view them.
# [http://proteopedia.org/wiki/index.php/Topic_pages Topic Pages] in Proteopedia, or its [http://proteopedia.org/wiki/index.php/Table_of_Contents Table of Contents].
# Random PDB Entry in Proteopedia (see ''Random'' at top left of this page in the ''navigation'' box).
# Search by molecule name or amino acid sequence at [http://www.pdb.org www.pdb.org], but remember that less than 10% of proteins have known structure. See also [[Practical Guide to Homology Modeling]] which includes instructions for finding empirical 3D models for a protein sequence.

==V. Explore Your Molecule==

===FirstGlance in Jmol===

The main tool we will use is '''FirstGlance in Jmol''': [http://firstglance.jmol.org FirstGlance.Jmol.Org]. (To google it later, use the single word (no space) '''firstglance'''.)

* Use '''Firefox''' (or Safari). The molecule will rotate slowly with jerky jumps in other browsers. Internet Explorer and Edge are especially bad for Jmol.

*Enter your 4-character PDB code at FirstGlance, and you should see the molecule you have chosen.

*Get familiar with what the ''molecule information tab'' tells you. Ask about anything you don't understand.

*In the ''Views'' tab, there are 10 links at the top that show you different aspects of the molecule. Try them all, as well as any of the other tools in FirstGlance that interest you.
<ul><ul>
FirstGlance does NOT use '''[[Java]]''' unless you tell it to. Using Java will make larger proteins load faster, rotate more smoothly, and change views quicker. Use '''Firefox or Internet Explorer''' (or Safari) for Java. Chrome and Edge do not support Java. In your Java-compatible browser, display a molecule in FirstGlance, and then click on the ''Preferences'' tab in FirstGlance. See [[Installing and enabling Java]].
</ul></ul>
Here are two Views in FirstGlance that will be used in your report:

====A. Hydrophobic/Polar====
*Water-soluble proteins have polar/charged amino acids nearly everywhere on their surfaces (Examples: small [[2hhd]], large [[1igy]]). Patches of hydrophobic amino acids on the surfaces of soluble proteins are usually less than ~10 å in their smaller diameter, and usually recessed.
*Hydrophobic surface patches may be buried in chain-to-chain contacts -- check the [[#Section_6:_Biological_Unit|biological unit]] (example: [[Lac_repressor#Structure_of_the_lac_repressor|lac repressor homodimer]]).
*Large, protruding hydrophobic surface areas (>25 Å in their smaller diameter) may indicate transmembrane proteins (insoluble). Examples:
**[http://firstglance.jmol.org/fg.htm?mol=7ahl 7ahl]
**showing [http://opm.phar.umich.edu/protein.php?pdbid=7ahl bilayer boundaries] (click on "Jmol"; ligand toggles boundaries).
**[http://firstglance.jmol.org/fg.htm?mol=1bl8 1bl8]
**[[Gramicidin Channel in Lipid Bilayer]].

====B. Charge====
Most proteins have roughly equal numbers of positive and negative charges intermixed on their surfaces. Surface patches of exclusively positive charge often bind nucleic acids (negatively charged because of their phosphates). For example, examine the protein surface charges where the gal4 transcriptional regulator binds DNA ([http://firstglance.jmol.org/fg.htm?mol=1d66 1d66]).

==VI. Report Slides==
Answer the questions below in slides, using [http://slides.google.com '''Google Slides'''] or powerpoint.

*Name your report '''YourLastName-IW2018''', for example Malvankar-IW2018. ''If the name of your report does not begin with your family name, you will lose 2 points.'' When completed, email a link to your report to '''TAs''' for grading.
::(While viewing your report Google slides, click the blue Share button at the upper right, then Get shareable link, and paste the link into the email.)
:You will ''not'' be asked to present your report in class.

*Each slide MUST be labeled at the top with its section number, e.g. ''Section 1''.

*Each Section below may be answered in a single slide, or multiple slides. For example, suppose you want to show two snapshots for ''Section 3'', and make separate comments. You may choose to use two slides, labeled ''Section 3A'' and ''Section 3B''.

*For full credit, your slides must include at least 2 animations.

*Additional work beyond the minimum required may earn extra credit.

*Due date: 22 October 2018 .

This is not a test. It is to help you learn by doing. Ask for help!
 
[https://drive.google.com/file/d/1EswWlN_X6qwwrkP7d16mLOR0d4-1fVAN/view?usp=sharing Example of a Completed Report] (You may import these slides into a new presentation of your own, and use them as a templates, putting in your own content.)

===Section 1: Identity===
*The label ''Section 1'' at the top (and so forth for every slide).
*Your name.
*Your major; grad students, give the name of your track (Micro, BQBS, MCGD, etc.) and whose lab you work in.
*Your [[PDB identification code]].
*The name of your molecule.
*The function of your molecule (briefly, one sentence).
*The experimental method and [[resolution]] (or number of models for NMR). Available in the '''molecule information tab''' in FirstGlance.

*A snapshot of your molecule.
<center><table border="0" width="80%" style="background:#ffffa0;"><tr><td><center>
[http://firstglance.jmol.org/slides.htm How to make a snapshot],
 
is also linked at the bottom left in ''FirstGlance''.
</center></td></tr></table></center>

===Section 2: Composition===
*The number of
**Protein chains ([[Chain|What does "chain" mean?]])
**DNA chains
**RNA chains
:Available in the '''molecule information tab''' in FirstGlance: Chain Details. You can identify a residue in any chain by touching it with the mouse (spinning off!). DNA residues are DA, DG, DC, DT while RNA residues are A, G, C and U.
*Ligands and Non-Standard Residues: Give the one to three-letter abbreviations and '''full names''' for all ligands and non-standard residues. If none, so state. ([[Standard residues]])
:The ''molecule information tab'' in FirstGlance lists the 1 to 3-letter abbreviations for each ligand and non-standard residue, and their full names. Click on an abbreviation to locate that entity in the model. See also ''Composition'' in FirstGlance's Views tab. 

===Section 3: Evolutionary Conservation===
See [[Introduction to Evolutionary Conservation]].   (Example 4d7b: [http://consurf.tau.ac.il Consurf], [http://firstglance.jmol.org/fg.htm?mol=http%3A//bioinformatics.org/molvis/atlas/pdb/4d7b_consurf1454623073_pipe.pdb Result])

Does your molecule have a highly conserved region? If so, what is its function? If there is no highly conserved region, is there a highly variable region? Show two snapshots illustrating a highly conserved region, and a contrasting region.

See [[How to see conserved regions]].

:If Proteopedia lacks a pre-calculated ''Evolutionary Conservation'' for your molecule, and you do your own calculation at the ConSurf Server, be sure to include the address of the ConSurf result in your report slide!.

===Section 4: Hydrophobic/Polar===
Do you think your molecule is water soluble? Support your conclusion with a snapshot. Be sure to use the ''Hydrophobic/Polar'' view from FirstGlance in a snapshot. Optionally, you may show other views in other snapshots.

===Section 5: Hydrophobic Core===
Are there hydrophobic cores in your molecule? For soluble proteins, expect a hydrophobic core in ''[http://proteopedia.org/wiki/index.php/Domain each domain]''. Support your conclusion with a snapshot. Be sure to use the ''Hydrophobic/Polar'' view from FirstGlance and turn on the ''Slab'' button.

===Section 6: Charge===

Are there any areas on the surface of your molecule with only positive (or negative) charges? Show snapshots illustrating your conclusions. Be sure to use the ''Charge'' view from FirstGlance in your snapshots.

===Section 7: Cation-Pi Interactions===

Use [http://capture.caltech.edu/ CaPTURE] program to identify energetically significant cation-pi interactions within proteins

Show a snapshot of an energetically significant [[Cation-pi_interactions|cation-pi interaction]]. Include a distance monitor in your snapshot. Also paste in the report from CaPTURE confirming its energetic significance. The cation-pi interaction tool, and instructions for measuring distances, are in the ''Tools Tab''.

===Section 8: Biological Unit===
In FirstGlance, in the ''molecule information tab'' click ''Biological Unit''. (It is also in the ''Resources Tab''.)

How many total polymer chains (protein + DNA + RNA) are in the asymmetric unit? The Biological unit?

Show side-by-side two snapshots comparing the asymmetric unit with the biological unit. The ''Cartoon'' representation in FirstGlance is best for these snapshots. Make sure to label which is which.

===Section 9 - Contacts/Non-covalent Bonds===
Example: [http://firstglance.jmol.org/fg.htm?mol=4d7b 4d7b].
#Click ''Contacts'' in the ''Tools Tab'' in FirstGlance.
#Change target selection to ''Residues/Groups''.
#Click on something small to select it as a "target", such as a ligand, or a single amino acid. Choose an amino acid with an uncharged polar side chain, such as Ser, Thr, Asn, Gln, Tyr, His.
#Click the link to ''Show atoms contacting target''.
#Click ''Center contacts''.
#Uncheck ''Backbones''.
#Click the 4th thumbnail image [[Image:Contact4.gif]] to display the contacts as balls and sticks colored by element. The element color key is at the bottom of the Contacts help panel in FirstGlance.
#Zoom in (and click ''Return to Contacts'' if necessary).
#Uncheck all categories of non-covalent bonds.
#Check ''hydrogen-bonded non-water''. (Review [[Hydrogen_bonds#Donor_and_Acceptor_Atoms|hydrogen bonds]].)
#Double click the hydrogen bond donor and acceptor atoms to insert a distance monitor.

Describe the moiety you selected as a target. Include a snapshot showing '''exactly one hydrogen bond'''. Be sure to identify the two entities (amino acids, nucleotides, ligand) by name, chain, and sequence number. I need enough detail to be able to reproduce what you are reporting.

===Section 10 - How Structure Supports Function===
Write a brief description ''in your own words'' (avoid plagiarism!) of how the structure of this protein supports its function. Doing some online research will strengthen your description.

Include links to supporting references. Wikipedia can be cited, but authoritative sources, such as peer-reviewed scientific journal articles or government websites, will have more weight.

Your description should be at least 75 words. More work, if well done, will earn more credit.

Section 10 in the [https://drive.google.com/file/d/1NyNhIJTVhmdNwry6Q3nxucig9x6e5-jx/view?usp=sharing Sample Report] is longer than the minimum required, but illustrates the sort of thing that could be done if you can spend the time.

==VII. See Also==
* [[User:Eric Martz/Introduction to Structural Bioinformatics]], a list of courses and workshops at various levels, including earlier versions of this segment.

==VIII. Notes and References==
<references />

Nikhil Malvankar:

* Full Real Name: Nikhil S. Malvankar
* Position: Postdoctoral Research Associate

* Institution (NO ABBREVIATIONS): University of Massachusetts, Amherst

* City, State/Province, Country: Amherst, MA, USA

* Field of Expertise or Study: Biophysics

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[[User:Nikhil Malvankar/Workbench/Geobacter pilus]]

User:Nikhil Malvankar

2014-07-18T23:33:25Z

Nikhil Malvankar: