3Dstructurevisualizationand

highqualityimaging.
Chimera

VincentZoete2008
Contact:vincent.zoete@isbsib.ch

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Table of Contents
PresentationofChimera.......................................................................................................................3
Exercise1.............................................................................................................................................4
LoadingastructureintoChimera....................................................................................................4
Moving/zooming............................................................................................................................5
Ribbonrepresentation......................................................................................................................5
Coloringthesecondarystructureelements.TheModelPanel........................................................6
Changingbondsandatomsdisplay.Selections...............................................................................7
Selectionsusingtheselectmenu.................................................................................................7
Changingbonddisplayandcolor................................................................................................8
Selectionsusingthecommandline.............................................................................................9
Changingribbonattributes.............................................................................................................11
Calculatingandshowinghydrogenbonds.....................................................................................11
Showingtheligandsurface............................................................................................................12
Labelresidues................................................................................................................................13
Savingimage..................................................................................................................................13
Savingthesessionstatus................................................................................................................14
Closingthesession.QuittingChimera..........................................................................................14
Restoringaprevioussession..........................................................................................................14
Exercise2...........................................................................................................................................15
Loadingthemacromolecularstructure..........................................................................................15
Showingthemolecularsurfaceoftheprotein...............................................................................15
Changethesurfacecolor................................................................................................................16
Changingbondsandatomsdisplay...............................................................................................16
Clippingtheprotein.......................................................................................................................16
Surfacecapping..............................................................................................................................18
Exercise3...........................................................................................................................................19
Openaprevioussession.................................................................................................................19
Usingthesideview........................................................................................................................19
ChangingtheLighting...................................................................................................................20
Goingfurther......................................................................................................................................21

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Presentation of Chimera
Thisintroductorycourseabout3Dstructurevisualizationandhighqualityimagingforpublication
willmakeuseofthefreeprogramChimera.Theofficialwebsiteofchimeracanbefoundatthe
followingaddress:http://www.cgl.ucsf.edu/chimera
HereisabriefdescriptionofChimeraanditsfeaturestakenfromthewebsite:
UCSFChimeraisahighlyextensible,interactivemolecularvisualization
andanalysissystem.Chimeracanreadmolecularstructuresandassociated
datainalargenumberofformats,displaythestructuresinavarietyof
representations,andgeneratehighqualityimagesandanimationssuitable
forpublicationandpresentation.Inaddition,Chimeraprovidestoolsto:
show density maps and analyze microscopy data; utilize symmetry
information for the display of higherorder structures; display multiple
sequencealignments,withcrosstalkbetweenthesequencesandstructures;
andenableanalysisofmoleculardynamicstrajectoriesanddockingresults.
Chimeraisdistributedwithfulldocumentationandanumberoftutorials,andcanbedownloaded
freeofchargeforacademic,government,nonprofit,andpersonaluse.Chimeraisavailablefor
several platforms, including Windows, MacOS X, and Linux. For more information, see the
Chimera web site. Chimera is developed and supported by the Resource for Biocomputing,
Visualization,andInformaticsandisfundedbytheNIHNationalCenterforResearchResources
(grantP41RR01081).
Theprogramcanbedownloadedatthefollowingaddress:
http://www.cgl.ucsf.edu/chimera/download.html
Whenusingchimera,oneshouldcitethisreference:
Pettersen, E.F., Goddard, T.D., Huang, C.C., Couch, G.S., Greenblatt, D.M., Meng, E.C., and
Ferrin,T.E."UCSFChimeraAVisualizationSystemforExploratoryResearchandAnalysis."J.
Comput.Chem.25(13):16051612(2004).

ThefollowingexerciseswillbeusedastutorialstointroducesomebasiccommandsofChimera.
TheHIV1proteaseincomplexwiththeA77003inhibitor(1HVIinPDB)willbeusedasan
example.Youarehighlyencouragedtotestthedifferentoptionsthatyouwillencounterinthe
differentmenus.
Generally,scientificjournalsrequireEPSorTIFFimages,witha300dotsperinchresolutionor
higher,andwithRGBcolors.YouwillseehowtoobtainsuchimagesusingChimera.

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Exercise 1
Loading a structure into Chimera
TwomethodscanbeusedtoloadastructureintoChimera.
1)Ifthestructurefileispresentintheuser'scomputer,choosethemenuitemFile/Open.Then
select1HVI.pdbandclickOpen.

Under Unix or Mac OSX, the structure (for instance 1HVI.pdb) can also be loaded using the
followingcommand:
>chimera1HVI.pdb

2)Thestructuremightalsobefetchedfromadatabase,whenavailable.Choosethemenuitem
File/FetchbyID.ChoosethePDBdatabankandtypethePDBcode1HVI.ThenclickFetch

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Oncethestructurehasbeenloaded,allmacromolecule'sbondsshouldshouldappearinthewire
representation,whichisChimera'sdefault.

Moving / zooming
The macromolecule can be rotated by clicking the left button and dragging the cursor over
Chimera's window. The translation is obtained similarly, but using the central mouse button.
Finally,onecanzoominandoutusingthemouserightbutton.

Rotate:pressleftbuttonanddrag
Translate:presscentralbuttonanddrag
Zoom:pressrightbuttonanddrag

Command line...
MostofChimerafunctionalitiescanbeaccessedbytypingagivencommandintheCommand
line,whichissituatedatthebottomoftheChimerawindow.Ifthecommandlineisnotpresent,
displayitusingFavorites/CommandLine.AlineprecededbyCommandwillappearonthe
lowerpartoftheprincipalwindow.Examplesofhowtousethecommandlinewillbegivenlater.

Ribbon representation
The secondary structure elements can be shown using the ribbon representation. This can be
accessedthroughtheActions/Ribbonsubmenu.
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OneconvenientfeatureofChimeraisitsabilitytodetachsubmenussothatonecanusethem
severaltimeswithouttheneedtogoagainthroughthedifferentmenus.Forinstance,choosethe
Actions/Ribbon menuitem,andclickonthedottedlineatthetopofthissubmenu.Thesub
menuwillbedetachedandfreelymovable.

Theribbonrepresentation ofthesecondarystructureelements canbeswitchedonbychoosing

showintheRibbonmenu.Threevariantsareavailable:flat,edgedandrounded.You
cantrythemallbyclickingthecorrespondingmenuitem.Selecttherepresentationyouprefer.

Important:typeksdsspinthecommandlinetorecalculatethesecondarystructureelementsof
theprotein. Thisusesanimplementation ofthe Kabschand Sanderalgorithm for defining the
secondarystructureofproteins.
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Coloring
the
secondary
elements. The Model Panel

structure

Theribboncanbecoloredaccordingtothesecondarystructureelement,i.e.strand,helixorloop.
OpentheModelPanelbychoosingtheTools/Depiction/ColorSecondaryStructuremenuitem.
This will open the Color Secondary Structure window. Check the Helix, Strand and
Other boxes. For each one, it is possible to modify the default color by clicking on the
correspondingcoloredsquaretoopentheColorEditorwindowandchangetheRGBcursors
positions.ClickontheApplybuttonoftheColorSecondaryStructurewindowtoapplythe
coloring.

Youcantryseveralcolorcombinationsandkeeptheoneyouprefer.Finally,youcanclosethe
Ribbon,ColorSecondaryStructureandColorEditorwindowsbyclickingtheClose
button,orthetoprightXicon.

Changing bonds
Selections

and

atoms

display.

Theobjectiveofthispartistodisplayonlytheligand,theactivesitewatermoleculeandresidues
25and50oftheHIV1protease.Otherresidueswillbehiddentoclarifythefigure.Thiswill
requiretoselectpartsofthestructureandapplythemdifferentrepresentationschemes.
There are three ways of selecting atoms with Chimera: using the Select menu, using the
CommandLine,andusingthemousetoselectatomsfromthescreen.Thelatterwillbedescribed
later.

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Selections using the select menu

Open the Select menu anddetached it by clicking onthe upper dotted line. Verify that the
SelectionModeisreplace.Otherwise,chooseit.
ThefirstfourmenuitemsallowtoselectpartofthestructureaccordingtotheChain(protein
chains,ligand,water),theChemistry(chemicalnatureoftheatomsorfunctionalgroups),the
Residue(residuenameortype)andtheStructure(backbone,sidechains,secondarystructure,
etc...).Youcantrytoselectdifferentpartsofthecomplex.Forinstance,youcanselecttheligand
bychoosingtheSelect/Chain/hetmenuitem.Youwillseethattheselectedpartsofthestructures
aresurroundedbygreenthinlines.
NotethatthisHIV1proteaseinhibitorisalsodefinedbyitsligandnatureoritsresiduename
(i.e., A77). Therefore, it can be selected also by Select/Structure/ligand or
Select/Residue/A77.

Youcantryselectingdifferentpartsofthecomplex:thewatermolecule,chainsAorBofthe
protein,theaspartateresidues,thestrands,etc...
Itispossibletoappendaselectionwithanewone.Forinstance,toselectalltheprotein,youcan
choose the menu items Select/Selection Mode/append, then Select/Chain/A and
Select/Chain/B.
Finally,youcanclearallselectionsusingSelect/ClearSelection.

Changing bond display and color

SelectthereplacemodeforselectionsusingSelect/SelectionMode/replace andselectthe
ligand.
OpentheActions/AtomsBondsmenuanddetachitbyclickingontheupperdottedline.Choose
successivelythestick,ballandstick,sphereandwiremenuitemsandseehowtheligand
isdisplayedineachcase.Finally,choosetheballandstickrepresentation.
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OpentheActions/Colormenuanddetacheditbyclickingontheupperdottedline.Youcan
eventuallyclosesomeotherwindowstosavespaceonthescreen.Checktheatoms/bondsboxso
thatthecolorchangingwillbeappliedonlytoatomsandbonds.Then,youcanselectacolorfrom
theleftcolumnthatwillbeusedforallligandatoms.Anotherpossibilityistocolorallatoms
accordingtotheiratomtypes.Thisisobtainedbyclickingbyelement.

Similarly,selectthewatermolecule,displayitinballandstickandcoloritaccordingtotheatom
types.

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Selections using the command line

Close the Select window using the upper right X icon. If it's not already done, open the
command line by choosing Favorites/Command line. A line preceded by Command will
appear on the lower part of the principal window. This line can be used to type and execute
commandsrelativetoselectionanddisplay,forinstance.Toexecuteacommand,oneneedstotype
itinthecommandlineandpressReturn.Theunionandintersectionselectionkeywordsarenoted
|and&,respectively.Thenegationsymbolis~.
Adetaileddescriptionofthedifferentselectionpossibilitiesusingthecommandlineareprovidedin
thequickreferenceguide.Hereisalimitedlistshowingsomepossibleselectionsbasedonour
particularstructurethatyoucouldtry:
select:selecteverything.
select:.A:selectchainAoftheprotein.
select:.A,.B:selectchainsAandBoftheprotein.
selectligand:selecttheligand.
select:A77:selecttheresiduenamedA77.Anotherwaytoselecttheligand.
select:HOH:selecttheresiduenamedHOH,i.e.theactivesitewatermoleculeinourcase.
select:HOH|ligand:selectboththewatermoleculeandtheligand.
select:25:selectallresiduesthatarenumbered25inthePDB.Oneinbothchains.
select:25@CA:selectatomCofresidues25.
select:25.A:selectresidue25ofchainA.
select:25,50:selectallresiduesthatarenumbered25and50inthePDB.
selectstrand:selectstrands.

Selectalltheproteinbytypingselect:.A,.Binthecommandline,andhidetheatomsandbonds
usingAction/Atomsbonds/hide.

Selectresidues25and50bytypingselect:25,50inthecommandline,andshowtheminball
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and stick representation, colored according to the atom type: Action/Atoms Bonds/show,
Action/AtomsBonds/ballandstick,Actions/Colors/byelement.

Changing ribbon attributes

Ascanbeseen,thebackboneatomsoftheproteinresiduesarenotshown,whilethebondlengths
betweenthebackbone(displayedasaribbon)andCatomsareartificiallytoolong.Thisisdueto
thedefaultbehaviorofChimerathathidesbackboneatomswhenribbonsaredisplayed.Thiscanbe
modified by changing some attributes of the structure. From the Model Panel, open the
attributeswindow,andchoosethefalsevalueforribbonhidesbackboneatoms.

Calculating and showing hydrogen bonds

Chimera can calculate and display the hydrogen bonds between selected atoms. To show the
hydrogenbondnetworkbetweentheligand,theactivesitewatermoleculeandtheIle50backbone
atoms,thefollowingsequenceofactionscanbeused.Selecttheactivesitewatermolecule(using
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the command line or the Select menu). Open the Tools/Structure Analysis/FindHBond
window.ChecktheOnlyfindHbondswithbox,sothatitwillcalculateonlyhydrogenbonds
involvingtheactivesitewatermolecule.Finally,clickontheApplybutton.

YoucanchangethehydrogenbondcolorandlinewidthfromtheHBondParameterswindow,
andclickApplytoapplythem.

Showing the ligand surface

Selecttheligand.Then,opentheSurfacewindowusingAction/Surface.Clickshow.You
cantrymodifyingthesurfacerepresentationtomeshanddot.Then,gobacktothesolid
representation.
The surface quality may be increased by changing the corresponding attribute. In the Model
Panelwindow,selecttheMSMSligandsurfaceintheleftlist.Thenclickattributesandchange
thevertexdensityattributeto10.PressEntertoapplythechange.

Thissurfacegivesagoodideaofthevolumeoccupiedbytheligand.However,italsohidesthe
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molecule. To correct this, it can be made transparent. In the Surface window, click on
transparencyandselect80%.

Label residues
SelectatomsCofresidues25bytypingselect:25@CBinthecommandline.Openanddetach
theLabelwindowbychoosingtheActions/Labelmenuitem.IntheLabelwindow,choose
residue/name+specifier.Youcantryothertypesoflabeling.Eventually,youcanchangethe
labelcolorusingtheColorwindow,checkingresiduelabelsandchoosingacolor.

Saving image
ClearallselectionsusingSelect/ClearSelection.Chooseanorientationandzoomthatprovidesa
satisfying point of view. Then, select the File/Save Image menu item. In the new window,
chooseanimageresolutionof300dotsperunit(dpi).SelectMaintaincurrentaspectratio,and
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enteranImagewidthof6inches.ClickSaveAs.Waitwhiletheimageiscalculated.Select
theformatofthefilethatwillbesaved.ScientificjournalsgenerallyacceptTIFFandEPSfiles.
SelectTIFFinthiscase.Finally,chooseandFilenameandclickSave.

ImagesaresavedasRGBfigures.

Saving the session status

TheChimerasession(theactualrepresentation)canbesavedforfutureuseormodifications.This
canbemadeusingtheFile/SaveSessionAs...menu.SelectafilenameandclickSave.The
savedfileisactuallyapythonscriptandwillhavethe.pyextension.

Closing the session. Quitting Chimera

ThesessioncanbeclosedusingtheFile/CloseSessionmenuitem.OnecanquitChimerawith
File/Quit.

Restoring a previous session

ApreviouslysavedsessioncanberestoredusingtheFile/RestoreSessionmenuitem.Selectthe
file(witha.pyor.pycextension)andclickOpen.

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Exercise 2
Loading the macromolecular structure
Loadthe1HVI.pdbstructurefileintoChimerausingoneofthetwomethodsseeninexercise1.

Showing the molecular surface of the

protein
First,hidethebondsandatomsoftheproteinchainsandkeeponlytheligandandactivesitewater
moleculevisible.Todoso,selectthetwoproteinchains,forexamplebytypingSelect:.A,.Bin
thecommandline.ThenopenanddetachtheAction/AtomsBondsmenuandchoosethehide
menuitem.

Besurethattheproteinisstillselected.Ifnecessary,orincaseofdoubt,selectitagainasdescribed
above. Then, open and detach the Action/Surface menu, and choose show. Open the
Favorites/Model panel menu, select the MSMS main surface in the left list, then click
attributesandchangethevertexdensityto10.Ifyourcomputeristooslow,itmightbe
necessarytoreducethevertexdensityto5.

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Change the surface color

Besurethattheproteinisstillselected.OpenanddetachtheActions/Colorsmenu.Checkthe
surfacesbox,andclickonthecoloryouwouldliketoapplytotheproteinsurface.Thefollowing
imagehasbeenobtainedusingthecornflowerbluecolor.

Changing bonds and atoms display

Selecttheligandandwatermolecules,thendisplaytheminballandstickrepresentation,colored
accordingtotheatomtype.

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Clipping the protein

Itispossibletocuttheproteinsurfaceintogetabetterviewoftheligandbindingmodeinsidethe
bindingsite.Todoso,openanddetachtheTools/Depictionmenu.ThenchoosethePermodel
Clipping menu item. InModel, select the MSMS main surface and check the Enable
clippingbox.Rotatethestructuretogetabetterviewoftheclippingeffect.

Itispossibletomanipulatetheclippingpositionusingthemouse.ChecktheAdjustclippingwith
mousebox.Aclick/dragofthemousecentralbuttonwillcontrolthetranslationoftheclipping
plane.Aclick/dragofthemouserightbuttonwillmodifyitsorientation.Notethatthiscancelany
possibilityofproteintranslationorzoomusingthemouse.Youcangetbacktothedefaultbehavior
ofthemousebyuncheckingtheAdjustclippingwithmousebox.
Youcanselectanddisplayresidues25and50oftheHIV1proteasebytypingselect:25,50in
thecommandline,andthendisplaytheminthestickrepresentation.Thenchangethepositionand

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orientationoftheclippingplanetohaveagoodviewoftheinteractionsbetweentheseresiduesand
theligand.

Youcanalsogetagoodviewoftheshapeofaburiedbindingsiteusingtheslabmode.InthePer
ModelClippingmenu,checktheUseslabmodewiththicknessandchoose6forthethickness.
This creates aprotein slabthatyoucanmanipulate usingthemousebycheckingthe Adjust
clippingwithmousebox.

UnchecktheUseslabmodewiththicknessbox.

Surface capping
Aclipped surfacemaybecapped.Todoso,choosetheSurfaceCappingmenuiteminthe
Depictionwindow.IntheSurfaceCappingwindow,checktheCapsurfaceatclipplanes
box.YoucanchangethecolorofthecappingplanebycheckingtheUsecapcolorboxand
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choosethecolorbyclickingthecoloredsquarenexttoit.

Thiscappingcanalsobeappliedwiththeslabmode.
Saveanimageasdescribedinthepreviousexercise,thensavethesessionandcloseit.

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Exercise 3
Open a previous session
Openthefinalstateofexercise2usingtheFile/RestoreSessionmenu.Removingallclippingand
capping,andchangethebackgroundcolortoblackifnecessary.

Using the side view

OpentheViewingmenubychoosingtheFavorites/Sideviewmenuitem.TheSideView
taboftheViewingwindowshouldbeactive.Otherwise,clickontheSideViewtab.Youwill
seeareducedviewofthestructureappearingintheViewingwindow.Theverticallinesshows
theclippingplanesthatdefinetheregionofspacedisplayedonthescreen.Thesquaregivesthe
viewer'seyeposition.Theredlinesshowthefieldofvision.
ClickingtheViewAllbuttonadjuststhescaleandclippingplanepositionssothattheviewwill
includeeverythingthatisdisplayed.Itispossibletomovethepositionoftheviewer'seyeand
clippingplanesusingthemouse.

Ifyoudrawtherearclippingplaneclosertotheprotein,youwillseetheeffectofthedepthcueing,
whichcausesregionsfartherfromtheviewertobeshaded.Thedepthcueingparameterscanbe
changedintheEffectstaboftheViewingwindow.

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Changing the Lighting

Itmightbeusefultochangethelighting(intensityanddirection)togetabetterviewofthesystem.
Select the Lighting tab in the Viewing window. This tab displays the light sources and
parameters.Thekeylightisthedominantbrightersourceoflight.Thefilllightgivesasecondary
source.Thesolidarrowsintherightviewallowtomanipulate thelighting directions withthe
mouse.Youcantrydifferentlightingdirectionstoseetheireffect.

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Exercise 4
Loading the macromolecular structures
Loadthe1HVI.pdbstructurefileintoChimerausingoneofthetwomethodsseeninexercise1.
Thenopenthe1HII.pdbstructurefile.ThesestructurefilescorrespondtotheHIV1andHIV2
proteases,respectively.

TheyarenowtwoactivemodelsmentionedinthelowerlineofthemainChimerawindow.Model0
isHIV1protease(1HVI),whereasmodel1isHIV2protease(1HII).Displaybothmoleculesusing
differentcolors.Forthis,eachmodelcanbeselectedsubsequentlybytypingselect#0andselect
#1inthecommandline.

Showing the proteins as ribbons

Selectallheteroatomsandwatermoleculesbytypingselect:.het|:HOHinthecommandline,
anddeletethecorrespondingatomsusingAction/Atomsbonds/delete.Onlythetwoproteinsare
remaining in Chimera. Displaytheminribbonrepresentation, asexplained inexercise 1.Hide
bondsforbothmolecules.Recalculatethesecondarystructureelementsbytypingksdsspinthe
commandline.

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Match the two molecules.

structural alignment

Obtain

OpenthestructuralalignmenttoolusingTools/StructureComparison/MatchMaker.Clickon
Specificchain(s)inreferencestructurewithspecificchain(s)inmatchstructure.Then,select
chainsAandBof1HVIasreferencestructures,andmatchthemwithchainsAandBof1HII,
respectively. Select the NeedlemanWunsch algorithm and click OK. This will perform a
matchingofthetwomolecules,basedontheirsequenceand3Dstructuresimilarities.

After a short moment, the two molecules should be superimposed. A new window, called
MultiAlignViewerwillalsoappear,showingthecorrespondingsequencealignments.
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Thepercentageofsequenceidentitybetweenthetwomoleculescanbeobtainedusingthemenu
Tools/Percentidentity...oftheMultiAlignViewerwindow.TheresultsappearintheChimera
commandline.

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ThesecondarystructureelementscanbehighlightedinthesequenceusingtheMultiAlignViewer
command:Structure/SecondaryStructure/showactual.Secondarystructureelementsappearin
greenforstrandsandyellowforhelices.

Isolatedresiduesorgroupsofresiduescanbeselectedbyclickingoverthesequenceorinthe
Conservation line of the MultiAlignViewer, allowing the analysis of the proteins structural
differencesoranalogies.

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Itispossibletocolorthestructuresaccordingtothepercentageofconservationinthesequence
alignment. For this, open the Structure/Render by Conservation menu of the
MultiAlignViewer,thenclickApplyusingthedefaultvaluesforthedifferentoptions.Inthe
presentcase,themostconservedregionsarecoloredinredandthelessconservedonesinblue.

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Going further
TheseexercisesgiveonlyaverylimitedoverviewofwhatChimeraisable.Youcanfindadetailed
documentation,aswellassometutorials,atthefollowingaddress:
http://www.cgl.ucsf.edu/chimera/docindex.html
Here are some examples of images produced using Chimera that were taken from the official
Website.
Slicedpotassiumchannel

ParameciumBursariaChlorellaVirus

DNAandNetropsin

BluetongueVirusandViralRNA

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