diff --git a/education/HADDOCK24/RNA-Pol-III-2024/index.md b/education/HADDOCK24/RNA-Pol-III-2024/index.md index 725534393..c464cefbb 100644 --- a/education/HADDOCK24/RNA-Pol-III-2024/index.md +++ b/education/HADDOCK24/RNA-Pol-III-2024/index.md @@ -6,6 +6,7 @@ tags: [MS, Cross-links, cryo-EM, Interaction, HADDOCK, DISVIS, PowerFit, RNA Pol image: feature: pages/banner_education-thin.jpg --- +This tutorial was last updated on 18-02-2026.
This tutorial consists of the following sections: * table of contents @@ -745,7 +746,7 @@ in the map and calculate a cross-correlation score for each of them. PowerFit is open-source and available for download from our [Github repository][link-powerfit]{:target="_blank"}. To facilitate its use, we have developed a [web portal][link-powerfit-web]{:target="_blank"} for it. -The server makes use of either local resources on our cluster, using the multi-core version of the software, or GPGPU-accelerated grid resources of the +The server makes use of either local resources on our cluster, using the multi-core version of the software, or GPU-accelerated grid resources of the [EGI](https://www.egi.eu){:target="_blank"} to speed up the calculations. It only requires a web browser to work and benefits from the latest developments in the software, based on a stable and tested workflow. Next to providing an automated workflow around PowerFit, the web server also summarizes and higlights the results in a single page including some additional postprocessing @@ -1050,7 +1051,7 @@ Another explanation could be conformational changes in the structures that are n
### Setting up the full docking run using the cryo-EM fitted and refined core and C82 domains -We will setup a docking run to model the full complex including some information about the C31 cross-links and the missing two first wHTH domains of C34. For this we will use the PowerFit/Chimera, HADDOCK-refined structure (Core+C82+C34wHTH3) which we have generated as starting point. We will fix those domains in their original positions for the initial rigid-body docking stage and dock the two missing C34 wHTH domains including fragments of C31 for which we have cross-links. +We will setup a docking run to model the full complex including some information about the C31 cross-links and the missing two first wHTH domains of C34. For this we will use the PowerFit/ChimeraX, HADDOCK-refined structure (Core+C82+C34wHTH3) which we have generated as starting point. We will fix those domains in their original positions for the initial rigid-body docking stage and dock the two missing C34 wHTH domains including fragments of C31 for which we have cross-links. Connect to the [HADDOCK2.4 interface](https://wenmr.science.uu.nl/haddock2.4/submit/1){:target="_blank"} of the HADDOCK web server. @@ -1303,7 +1304,7 @@ ranking cluster, `clusterX_1.pdb`. In the ChimeraX command window type: -color by chain
+color bychain
hide atoms
show cartoon
distance /B:50@CB /F:91@CB
@@ -1564,7 +1565,7 @@ our [HADDOCK forum](https://ask.bioexcel.eu/c/haddock){:target="_blank"} hosted [link-haddock-register]: https://wenmr.science.uu.nl/auth/register/"HADDOCK web server registration" [link-molprobity]: http://molprobity.biochem.duke.edu "MolProbity" [link-powerfit]: https://github.com/haddocking/powerfit "PowerFit" -[link-powerfit-web]: https://alcazar.science.uu.nl/services/POWERFIT/ "PowerFit web server" +[link-powerfit-web]: https://wenmr.science.uu.nl/powerfit "PowerFit web server" [link-powerfit-register]: https://wenmr.science.uu.nl/auth/register "PowerFit registration" -[link-powerfit-submit]: https://alcazar.science.uu.nl/cgi/services/POWERFIT/powerfit/submit "PowerFit submission" -[link-powerfit-help]: https://alcazar.science.uu.nl/cgi/services/POWERFIT/powerfit/help "PowerFit submission" +[link-powerfit-submit]: https://wenmr.science.uu.nl/powerfit "PowerFit submission" +[link-powerfit-help]: https://www.bonvinlab.org/powerfit/manual.html "PowerFit help" diff --git a/education/Others/index.md b/education/Others/index.md index b994d851e..62aba45df 100644 --- a/education/Others/index.md +++ b/education/Others/index.md @@ -14,7 +14,7 @@ In this page you can find links and tutorials to two HADDOCK-complementary softw * [**PowerFit web server tutorial**](/education/Others/powerfit-webserver/): A small introduction into PowerFit to automatically place a high-resolution atomic structure in a - lower-resolution cryo-electron microscopy density map making use of our [web portal](https://alcazar.science.uu.nl/cgi/services/POWERFIT/powerfit) + lower-resolution cryo-electron microscopy density map making use of our [web portal](https://wenmr.science.uu.nl/powerfit) (does not require Linux). * [**DisVis web server tutorial**](/education/Others/disvis-webserver/): A small introduction into diff --git a/education/Others/powerfit-webserver/index.md b/education/Others/powerfit-webserver/index.md index 6521713a1..71fc2f771 100644 --- a/education/Others/powerfit-webserver/index.md +++ b/education/Others/powerfit-webserver/index.md @@ -2,10 +2,12 @@ layout: page title: "PowerFit web server Tutorial" excerpt: "A small tutorial on PowerFit web server for automatic rigid body fitting" -tags: [PowerFit, Cryo-EM, HADDOCK, Ribosome, Chimera, rigid body fitting] +tags: [PowerFit, Cryo-EM, HADDOCK, Ribosome, ChimeraX, rigid body fitting] image: feature: pages/banner_education-thin.jpg --- +This tutorial was last updated on 18-02-2026 and is up-to-date +with release v4.0.4.
This tutorial consists of the following sections: * table of contents @@ -20,12 +22,11 @@ It is open-source and available for download from our [Github repository][link-p To facilitate its use, we have developed a [web portal][link-powerfit-web]{:target="_blank"} for it. This tutorial demonstrates the use of the PowerFit web server. The server makes use of either -local resources on our cluster, using the multi-core version of the software, or GPGPU-accelerated grid resources of the +local resources on our cluster, using the multi-core version of the software, or GPU-accelerated grid resources of the [EGI](https://www.egi.eu){:target="_blank"} to speed up the calculations. It only requires a web browser to work and benefits from the latest developments in the software based on a stable and tested workflow. Next to providing an automated workflow around -PowerFit, the web server also summarizes and higlights the results in a single page including some additional postprocessing -of the PowerFit output using [UCSF Chimera][link-chimera]{:target="_blank"}. -Chimera is a visualization software and popular tool in the cryo-EM community for its volume visualization capabilities. +PowerFit, the web server also summarizes and higlights the results in a single page including some visualization of the +PowerFit output using [MolViewSpec][link-molviewspec]{:target="_blank"}. The case we will be investigating is a complex between the 30S maturing *E. coli* ribosome and RsgA, a GTPase. There are models ([2YKR][link-pdb]{:target="_blank"}) and a cryo-EM density map of around 9.8Å resolution @@ -55,19 +56,20 @@ _Structure._ *23*, 949-960 (2015). Throughout the tutorial, coloured text will be used to refer to questions, -instructions, and Chimera commands. +instructions, and ChimeraX commands. This is a question prompt: try answering it! This an instruction prompt: follow it! -This is a Chimera prompt: write this in the -Chimera command line prompt! +This is a ChimeraX prompt: write this in the +ChimeraX command line prompt! ## Setup/Requirements -In order to follow this tutorial you only need a **web browser**, a **text editor**, and [**UCSF Chimera**][link-chimera]{:target="_blank"} +In order to follow this tutorial you only need a **web browser**, a **text editor**, and [**UCSF ChimeraX**][link-chimerax]{:target="_blank"} (freely available for most operating systems) on your computer in order to visualise the input and output data. +ChimeraX is a visualization software and popular tool in the cryo-EM community for its volume visualization capabilities. Further, the required data to run this tutorial should be downloaded [**here**][link-data]{:target="_blank"}. Once downloaded, make sure to unpack the archive. @@ -77,22 +79,20 @@ Once downloaded, make sure to unpack the archive. Let us first inspect the data we have available, namely the cryo-EM density map and the structures we will attempt to fit. -Using Chimera, we can easily visualize and inspect the density and models, +Using ChimeraX, we can easily visualize and inspect the density and models, mostly through a few mouse clicks. -For this open the density map `ribosome-RsgA.map` and the PDB file of the ribosome which is already fitted into the map `ribosome.pdb`. +Open first the density map `ribosome-RsgA.map` and then the PDB file of the ribosome which is already fitted into the map `ribosome.pdb`. +ChimeraX will automatically guess their type. - UCSF Chimera Menu → File → Open... → Select the file + UCSF ChimeraX Menu → File → Open... → Select the file -Repeat this for each file. Chimera will automatically guess their type. - - -If you want to use the Chimera command-line instead, you need to first display it: +You can also use the ChimeraX command-line, however you might need to display it first: - UCSF Chimera Menu → Favorites → Command Line + UCSF ChimeraX Menu → Tools → Command Line and type: @@ -108,30 +108,18 @@ and type: In the `Volume Viewer` window, the middle slide bar provides control on the value at which the isosurface of the density is shown. At high values, the envelope will shrink while lower values might even display the noise in the map. -In the same window, you can click on `Center` to center the view on all visible molecules and the density. We will first make the density transparent, in order to be able to see the fitted structure inside: - - Within the Volume Viewer window click on the gray box next to Color - - -This opens the`Color Editor` window. - - -Check the Opacity box. - - -An extra slider bar appears in the box called A, for the alpha channel. - - -Set the alpha channel value to around 0.6. + + transparency #1 60 Notice that the density becomes transparent providing a better view of the fit of the ribosome model. On closer inspection, you can also discern a region of the density that is not accounted for by the ribosome structure alone: This should be the -binding location of RsgA. Although you could try and manually place the crystal +binding location of RsgA. If you cannot find it you can try to lower the value of the isosurface using the slider in the `Volume Viewer` window. +Although you could try and manually place the crystal structure in that region, finding the correct orientation is not straightforward. PowerFit can help you here as it will exhaustively sample all possible translations and rotations in order to find the best fit, based on an objective score. @@ -145,7 +133,7 @@ the three translational and three rotational degrees of freedom. In short, PowerFit will try to systematically fit the structure in different orientations at every position in the map and calculate a cross-correlation score for each of them. -In order to perform the search PowerFit requires three different things: +In order to perform the search, PowerFit requires three different things: a high-resolution atomic structure of the biomolecule to be fitted (`RsgA.pdb`), a target cryo-EM density map to fit the structure in (`ribosome-RsgA.map`), and the resolution, in Ångstrom, of the @@ -153,7 +141,7 @@ density map (`9.8`). This is also the minimal required input for the web server To run PowerFit, go to -https://alcazar.science.uu.nl/services/POWERFIT +https://wenmr.science.uu.nl/powerfit On this page, you will find the most relevant information about the server as well as the links to the local and grid versions of the portal's submission page. @@ -161,8 +149,8 @@ On this page, you will find the most relevant information about the server as we [Register][link-powerfit-register]{:target="_blank"} for getting access to the webserver (or use the credentials provided in case of a workshop). -You can click on the "**Register**" menu from any PowerFit page and fill the required information. Registration is not automatic but is usually processed within 12h, so be patient. +When you are registered you can use all the software provided in the webserver. ### Step2: Define the input files and parameters and submit @@ -170,7 +158,7 @@ Click on the "**Submit**" menu to access the [input form][link-powerfit-submit]{
-
+ Complete the form by filling the required fields and selecting the respective files (most browsers should also support dragging the files onto the selection button): @@ -192,9 +180,7 @@ pre- and post-processing steps might substantially increase the time until the r While the calculations are running, open a second tab and go to -https://alcazar.science.uu.nl/services/POWERFIT - -Then click on the "**Help/Manual**" menu. +https://www.bonvinlab.org/powerfit/manual.html Here, you can have a look at the several features and options of PowerFit and read about the meaning of the various input parameters (including the ones under "**Advanced options**"). @@ -205,32 +191,25 @@ sampled. Lower values will cause PowerFit to perform a finer search, at the expense of increased computational time. The default value is 10°, but it can be lowered to 5° for more sensitive searches, or raised to 20° if time is an issue or if there aren't sufficient computational resources. -The number of processors used for the calculation is fixed on the web server side to 8 processors. -This number can of course be changed when using the local version of PowerFit. - ## Analysing the results Once your job has completed, and provided you did not close the status page, you will be automatically redirected to the results -page (you will also receive an email notification). The results page presents a summary split into several sections: +page (you will also receive an email notification). -* `Status`: In this section you will find a link from which you can download the output data as well as some information -about how to cite the use of the portal. -* `Solutions`: The best 15 non-redundant solutions found, ordered by their -cross correlation score. The first column shows the rank, column 2 the cross correlation -score, column 3 and 4 the Fisher z-score and the number of standard -deviations. The table is created with values taken from the file `solutions.out`. -* `Fit N`: Summary of the previous table for the 10 best fitted structures according to the cross correlation score. -A PDB of the solution can be downloaded and 6 images of the PDB within the density map are shown, covering different -views over the scene. +On the interactive results page you can look at the top + +
+ +
-If you don't' want to wait for your run to complete, you can access the precalculated results of a run submitted +If you don't want to wait for your run to complete, you can access the precalculated results of a run submitted with the same input [here][link-powerfit-tutorial]{:target="_blank"}. The higher the cross-correlation score the better the fit. But also important is the Fisher z-score (the higher the better), which, together with its associated number of standard deviations (σ difference), is an excellent indicator of the accuracy of a fit (see for details [van Zundert and Bonvin, J. Struct. Biol. (2016)](https://doi.org/10.1016/j.jsb.2016.06.011){:target="_blank"} -and PowerFit [help page](link-powerfit-help){:target="_blank"}). +and PowerFit [help page][link-powerfit-help]{:target="_blank"}). To enhance the interpretation of the results in the `Solutions` table, the entries are colored in a green gradient up to a sigma difference of 3. @@ -261,11 +240,11 @@ likely location of the center of mass of the structure. with date and timing information. -Let us now inspect the solutions in Chimera. +Let us now inspect the solutions in ChimeraX. Open the density map, the *lcc.mrc* cross-correlation map, and the 10 -best-ranked solutions in Chimera. +best-ranked solutions in ChimeraX. Use for this either the `Menus` or the `command line interface` as explained [before](#inspecting-the-data), e.g.: @@ -284,8 +263,13 @@ Use for this either the `Menus` or the `command line interface` as explained [be -Make the density map transparent again, by adjusting the alpha channel value to -0.6. The values of the `lcc.mrc` slider bar correspond to the cross-correlation +Make the density map transparent again. + + + transparency #1 60 + + +The values of the `lcc.mrc` slider bar correspond to the cross-correlation score found. In this way, you can selectively visualize regions of high or low cross-correlation values: i.e., pushing the slider to the right (higher cutoff) shows only regions of the grid with high cross-correlation scores. @@ -294,15 +278,16 @@ As you can see, PowerFit found quite some local optima, one of which stands out (if the rotational search was tight enough). Further, the 10 best-ranked solutions are centered on regions corresponding to local cross-correlation maxima. -To view each fitted solution individually, open the `Model Panel` window. +To view each fitted solution individually, open the `Model Panel` window if it is +not already open -Favorites → Model Panel +Tools → Models The window shows each model -and its associated color that Chimera has processed. To show or hide a specific -model you can click the box in the `S` column. +and its associated color that ChimeraX has processed. To show or hide a specific +model you can click the box in the column with an eye. Go through the 10 solutions one by one to asses their goodness-of-fit @@ -314,7 +299,7 @@ model you can click the box in the `S` column. - In a new Chimera session, reopen the density map and the fit that you find + In a new ChimeraX session, reopen the density map and the fit that you find best. @@ -334,23 +319,27 @@ You now have combined the ribosome structure with the rigid-body fit of RsgA calculated by PowerFit, yielding an initial model of the complex. Take a closer look at residues `R47` to `H51` which are contributing to the interface with the ribosome. -In the command line of Chimera, type the +In the command line of ChimeraX, type the following instructions to center your view on these residues and highlight their interactions: - show #2:47-51 zr<5 & #1 || #2:47-51 - focus #2:47-51 zr<5 & #1 || #2:47-51 + hide #2 atoms
+ show #2 cartoons
+ sel #3:47-51
+ view sel
+ contacts sel distanceOnly 5.0 makePseudobonds true reveal true
Take some time to inspect the model, paying particular attention to these five residues and their spatial neighbors. - Are there any clashes between the ribosome and RsgA chains? + Are there any clashes between the ribosome and RsgA chains? Show the selection as spheres to visualize this better. -Chimera also includes a tool to locally optimize the fit of a rigid structure + +ChimeraX also includes a tool to locally optimize the fit of a rigid structure against a given density map, which can be an additional help on top of the PowerFit calculations. Make the main display window active by clicking on it, @@ -366,14 +355,14 @@ Press Fit to start the optimization. - Does the Chimera local fit optimization tool improve the results of PowerFit? + Does the ChimeraX local fit optimization tool improve the results of PowerFit? -The scoring function used by Chimera to estimate the quality of the fit makes +The scoring function used by ChimeraX to estimate the quality of the fit makes our model worse, increasing the number of clashes between the ribosomal RNA and RsgA. Click `Undo` in the `Fit in Map` window to undo the optimization. -Next, we will try to optimize the fit using the cross-correlation that Chimera +Next, we will try to optimize the fit using the cross-correlation that ChimeraX provides. @@ -414,14 +403,15 @@ for computational biomolecular research. [link-powerfit]: https://github.com/haddocking/powerfit "PowerFit" -[link-powerfit-web]: https://alcazar.science.uu.nl/services/POWERFIT/ "PowerFit web server" -[link-powerfit-register]: https://wenmr.science.uu.nl/auth/register/powerfit"PowerFit registration" -[link-powerfit-submit]: https://alcazar.science.uu.nl/cgi/services/POWERFIT/powerfit/submit "PowerFit submission" -[link-powerfit-help]: https://alcazar.science.uu.nl/cgi/services/POWERFIT/powerfit/help "PowerFit submission" -[link-chimera]: https://www.cgl.ucsf.edu/chimera/ "UCSF Chimera" +[link-powerfit-web]: https://wenmr.science.uu.nl/powerfit "PowerFit web server" +[link-powerfit-register]: https://wenmr.science.uu.nl/auth "PowerFit registration" +[link-powerfit-submit]: https://wenmr.science.uu.nl/powerfit "PowerFit submission" +[link-powerfit-help]: https://www.bonvinlab.org/powerfit/manual.html "PowerFit submission" +[link-chimerax]: https://www.cgl.ucsf.edu/chimerax/ "UCSF ChimeraX" [link-data]: https://alcazar.science.uu.nl/cgi/services/POWERFIT/powerfit/powerfit-tutorial.tgz "PowerFit tutorial data" [link-density]: https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-1884 "Ribosome RsgA density" [link-pdb]: https://www.rcsb.org/pdb/explore/explore.do?structureId=2YKR "PDBid 2YKR" [link-haddock-tuto]: https://bonvinlab.org/education/Others/powerfit#HADDOCK-cryoEM "HADDOCK with cryoEM data" [link-powerfit-tuto]: https://bonvinlab.org/education/Others/powerfit "Powerfit command-line tutorial" [link-powerfit-tutorial]: https://alcazar.science.uu.nl/cgi/services/POWERFIT/powerfit/example "Powerfit tutorial results page" +[link-molviewspec]: https://molstar.org/mol-view-spec/ "molviewspec" \ No newline at end of file diff --git a/education/Others/powerfit-webserver/powerfit_results.png b/education/Others/powerfit-webserver/powerfit_results.png deleted file mode 100644 index 06e2c7200..000000000 Binary files a/education/Others/powerfit-webserver/powerfit_results.png and /dev/null differ diff --git a/education/Others/powerfit-webserver/powerfit_submission.png b/education/Others/powerfit-webserver/powerfit_submission.png index 24d6f5ad1..2ad6a5acc 100644 Binary files a/education/Others/powerfit-webserver/powerfit_submission.png and b/education/Others/powerfit-webserver/powerfit_submission.png differ diff --git a/education/Others/powerfit-webserver/webservice_result_page.png b/education/Others/powerfit-webserver/webservice_result_page.png new file mode 100644 index 000000000..0d0b7b129 Binary files /dev/null and b/education/Others/powerfit-webserver/webservice_result_page.png differ diff --git a/education/Others/powerfit/index.md b/education/Others/powerfit/index.md index 3253edde4..99061fac2 100644 --- a/education/Others/powerfit/index.md +++ b/education/Others/powerfit/index.md @@ -2,10 +2,12 @@ layout: page title: "PowerFit Tutorial" excerpt: "A small tutorial on PowerFit for automatic rigid body fitting" -tags: [PowerFit, Cryo-EM, HADDOCK, Ribosome, Chimera, rigid body fitting] +tags: [PowerFit, Cryo-EM, HADDOCK, Ribosome, ChimeraX, rigid body fitting] image: feature: pages/banner_education-thin.jpg --- +This tutorial was last updated on 18-02-2026 and is up-to-date +with release v4.0.4.
This tutorial consists of the following sections: * table of contents @@ -18,11 +20,12 @@ This tutorial consists of the following sections: PowerFit is a software application developed to fit atomic resolution structures of biomolecules to cryo-electron microscopy (cryo-EM) density maps. -It is open-source and available for download on [Github][link-powerfit]. +It is open-source and available for download on [Github][link-powerfit]. In addition +you can find a manual on usage of PowerFit [here][link-manual]. This tutorial will show you how to utilize PowerFit by applying it to an E.coli ribosome case. To follow this tutorial, you need, in addition to PowerFit, the -[UCSF Chimera][link-chimera] visualization software, a popular tool in the +[UCSF ChimeraX][link-chimerax] visualization software, a popular tool in the cryo-electron microscopy community for its volume visualization capabilities. We will further discuss the limits of rigid body fitting, and how HADDOCK can alleviate some of the shortcomings. We provide the data necessary to run this @@ -41,13 +44,13 @@ _AIMS Biophysics_. *2*, 73-87 (2015). _Structure._ *23*, 949-960 (2015). Throughout the tutorial, colored text will be used to refer to questions or -instructions, Linux and/or Chimera commands. +instructions, Linux and/or ChimeraX commands. This is a question prompt: try answering it! This an instruction prompt: follow it! -This is a Chimera prompt: write this in the -Chimera command line prompt! +This is a ChimeraX prompt: write this in the +ChimeraX command line! This is a Linux prompt: insert the commands in the terminal! @@ -74,7 +77,7 @@ ribosome has already been properly fitted in the density. *In case you might run this tutorial on your own*, make sure to have the required -software installed ([UCSF Chimera][link-chimera] and [PowerFit][link-powerfit]), +software installed ([UCSF ChimeraX][link-chimerax] and [PowerFit][link-powerfit]), and download the data to run this tutorial from our GitHub data repository [here][link-data] or clone it from the command line @@ -89,14 +92,14 @@ data repository [here][link-data] or clone it from the command line Let us first inspect the data we have available, namely the cryo-EM density map and the structures we will attempt to fit. -Using Chimera, we can easily visualize and inspect the density and models, +Using ChimeraX, we can easily visualize and inspect the density and models, mostly through a few mouse clicks. Open the density map together with the ribosome and KsgA. - chimera ribosome-KsgA.map ribosome.pdb KsgA.pdb + chimerax ribosome-KsgA.map ribosome.pdb KsgA.pdb In the `Volume Viewer` window, the middle slide bar provides control on the @@ -104,11 +107,9 @@ value at which the isosurface of the density is shown. At high values, the envelope will sink while lower values might even display the noise in the map. We will first make the density transparent, to see the fitted structure inside: -* Within the `Volume Viewer` click on the gray box next to `Color`, which opens - the `Color Editor` window. -* In there, check the `Opacity` box. An extra slider bar appears in the box - called `A`, for the alpha channel. -* Set the alpha channel value to around 0.6. + + transparency #1 60 + Notice that the density becomes transparent providing a better view of the fit of the ribosome model. On closer inspection, you can also discern a region of @@ -133,7 +134,7 @@ on the map and calculate a cross-correlation score for each of them. map. - powerfit ribosome-KsgA.map 13 KsgA.pdb -d run-KsgA -a 20 -p 2 -l + powerfit ribosome-KsgA.map 13 KsgA.pdb -d run-KsgA -a 20 -p 2 While performing the search, PowerFit will update you on the progress of the @@ -161,10 +162,8 @@ option specifies where the results will be stored while the `-p` option specifies the number of processors that PowerFit can use during the search, to leverage available CPU resources. -Finally, the `-l` flag applies a Laplace pre-filter on the density data, which -increases the cross-correlation sensitivity by enhancing edges in the density. In this example scenario, all other options are left at their default values -but feel free to explore them. +but feel free to explore [them][link-manual]. ## Analyzing the results @@ -186,14 +185,14 @@ with date and timing information. Open the density map, the *lcc.mrc* cross-correlation map, and the 10 -best-ranked solutions in Chimera. +best-ranked solutions in ChimeraX. - chimera ribosome-KsgA.map run-KsgA/lcc.mrc ribosome.pdb run-KsgA/fit_*.pdb + chimerax ribosome-KsgA.map run-KsgA/lcc.mrc ribosome.pdb run-KsgA/fit_*.pdb -Make the density map transparent again, by adjusting the alpha channel value to -0.6. The values of the `lcc.mrc` slider bar correspond to the cross-correlation +Make the density map transparent again to 60. +The values of the `lcc.mrc` slider bar correspond to the cross-correlation score found. In this way, you can selectively visualize regions of high or low cross-correlation values: i.e., pushing the slider to the right (higher cutoff) shows only regions of the grid with high cross-correlation scores. @@ -202,10 +201,10 @@ As you can see, PowerFit found quite some local optima, one of which stands out (if the rotational search was tight enough). Further, the 10 best-ranked solutions are centered on regions corresponding to local cross-correlation maxima. -To view each fitted solution individually, in the main panel, go to `Favorites` -→ `Model Panel` to open the `Model Panel` window. The window shows each model -and its associated color that Chimera has processed. To show or hide a specific -model you can click the box in the `S` column. +To view each fitted solution individually, in the main panel, go to `Tools` +→ `Models` to open the `Model Panel` window. The window shows each model +and its associated color that ChimeraX has processed. To show or hide a specific +model you can click the box in the column with an eye. Go through the 10 solutions one by one to appreciate their goodness-of-fit @@ -214,12 +213,13 @@ model you can click the box in the `S` column. Do you agree with what PowerFit proposes as the best solution? + - In a new Chimera session, reopen the density map and the fit that you find + In a new ChimeraX session, reopen the density map and the fit that you find best. Replace *?* by the appropriate solution number. - chimera ribosome-KsgA.map ribosome.pdb run-KsgA/fit_?.pdb + chimerax ribosome-KsgA.map ribosome.pdb run-KsgA/fit_?.pdb You now have combined the ribosome structure with the rigid-body fit of KsgA @@ -228,29 +228,30 @@ experiments performed on this complex indicate three charged residues of KsgA - `R221`, `R222`, and `K223` - that are of special importance for the interaction. -In the same session of Chimera where you have your chosen fitted KsgA -structure, go to `Favorites` → `Command Line`. A command line is now present -below the main viewing window. In the command line of Chimera, type the +In the same session of ChimeraX where you have your chosen fitted KsgA +structure, typ in the command line of ChimeraX the following instructions to center your view on these residues and highlight their interactions: - show #2:221-223 zr<5 & #1 || #2:221-223 - center #2:221-223 zr<5 & #1 || #2:221-223 + sel #3:221-223
+ view sel
+ contacts sel distanceOnly 5.0 makePseudobonds true reveal true
+ Take some time to inspect the model, paying particular attention to these three residues and their spatial neighbors. - Are there any clashes between the ribosome and KsgA chains? + Are there any clashes between the ribosome and KsgA chains? Show the selection as spheres to visualize this better. Is the mutagenesis data explained by the model, i.e. are the three charged amino acids involved in strong interactions? -Chimera also includes a tool to locally optimize the fit of a rigid structure +ChimeraX also includes a tool to locally optimize the fit of a rigid structure against a given density map, which can be an additional help on top of the PowerFit calculations. Make the main display window active by clicking on it, then go to `Tools` → `Volume data` → `Fit in Map`. In the newly opened `Fit in @@ -259,14 +260,14 @@ Map` window, select the best-fitted structure of PowerFit (`fit_?.pdb`) as Press `Fit` to start the optimization. - Does the Chimera local fit optimization tool improve the results of PowerFit? + Does the ChimeraX local fit optimization tool improve the results of PowerFit? -The scoring function used by Chimera to estimate the quality of the fit makes +The scoring function used by ChimeraX to estimate the quality of the fit makes our model worse, increasing the number of clashes between the ribosomal RNA and KsgA. Click `Undo` in the `Fit in Map` window to undo the optimization. -Next, we will try to optimize the fit using the cross-correlation that Chimera +Next, we will try to optimize the fit using the cross-correlation that ChimeraX provides. Click `Options` and check the `Use map simulated from atoms, resolution` box and fill in `13` for resolution. Check the `correlation` radio button and uncheck the `Use only data above contour level from first map`. @@ -304,10 +305,10 @@ HADDOCK model, generated by combining the cryo-EM map, the PowerFit centroid positions, and the mutagenesis data, in the tutorial data folder. - Open the density map in Chimera and load the best-ranked HADDOCK model. + Open the density map in ChimeraX and load the best-ranked HADDOCK model. - chimera ribosome-KsgA.map HADDOCK-ribosome.pdb HADDOCK-KsgA.pdb + chimerax ribosome-KsgA.map HADDOCK-ribosome.pdb HADDOCK-KsgA.pdb Does HADDOCK improve the quality of the model, i.e. are the number of clashes @@ -367,6 +368,7 @@ suggestions, feel free to contact us via email or by submitting an issue in the appropriate Github repository. [link-powerfit]: https://github.com/haddocking/powerfit "PowerFit" -[link-chimera]: https://www.cgl.ucsf.edu/chimera/ "UCSF Chimera" +[link-chimerax]: https://www.cgl.ucsf.edu/chimerax/ "UCSF ChimeraX" [link-data]: https://github.com/haddocking/powerfit-tutorial "PowerFit tutorial data" [link-density]: https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-2017 "Ribosome KsgA density" +[link-manual]: https://bonvinlab.org/powerfit/manual.html "PowerFit manual" \ No newline at end of file