Work through the tour by clicking the green scene triggers.
When you reach the bottom of this textbox, scroll to continue.
When not in a scene animation, the center structure window is fully interactive. For example, you can click and drag to adjust the view of the molecules.
Below is a legend for the main chain colors; colors are styled to match the article.

LEGEND:
◼ Rpr1 RNA		◼ RPP1a (arm)
◼ pre-tRNA		◼ Pop5p (arm)
◼ Pop1p (head)		◼ RPP1b (arm)
◼ Pop6p (head)		◼ Pop8p (arm)
◼ Pop7pA (head)		◼ Pop4p (arm)
		◼ Rpr2p (arm)
		◼ Pop3p (arm)

For additional convenience, using the `Legend` toggle below the structure window, a legend can be toggled on at any time within the structure window.
Other convenience buttons can be found on the left panel.
For example, there is a 'high quality' toggle that can be triggered; however, it slows interaction substantially, and so you'll want to toggle it back off to manipulate the structure view in any way. Toggles triggered during an animated scene get queued to run AFTER the animations.

Because of the complexity of the structure and representations in various scenes, you may find 'artifacts' left over from prior scenes arise when viewing items out of order. If that looks to be happening, you can hit the 'initial-like' button in the second row below the structure window to restore the the initial settings before trying the scene again to see if it improves.

The RNAse P Cryo-EM structure

Brief overview of the structure (OPTIONAL)

Having the surface of all the proteins displayed is the way the image in the paper starts off, and they really serve to show the intimate interactions of the chains. However, those take a substantial amount of time to calculate or load, and so those will be opt-in, other than the pre-tRNA surface. So give things a moment, once you click the next scene link. Load protein surfaces. However, the surfaces can make interaction with the structure sluggish and can also obscure other features, and so a good compromise is to leave the surfaces off for the most part and use the 'cartoon ribbon' view most of the time. Hide protein surfaces. Once they are loaded, turning the molecular surfaces off and on is immediate. (You can always use the toggles available to turn on specific ones, or click here to re-show all the protein surfaces). This is much faster than triggering 'load surfaces' again.

 

Similarly, loading Rpr1 RNA can take a moment...

 

Three metal ions are observed in the structure. Two magnesiums and one zinc. The ions are shown here with the proteins as backbones only to make the spheres representing the ions easy to spot. The bright green magnesiums highlight the active site where the 5' leader of the bound pre-tRNA is cleaved.
The first three nucleotides of the mature 5'-end of the tRNA are highlighted in red.

 

Overview of Rpr2p in the structure

Rpr2p is specific to yeast RNase P. All the other protein subunits known to be the yeast RNase P complex (& observed in the cryo-EM structure) of RNase P are also found in the related complex, yeast RNase MRP. Thus, Rpr2p's structure and function is expected to contribute specificity of the main RNA component of RNase P and interaction with the substrate.

 

In preparation for this section, the surfaces of the three proteins involved need to be loaded if the overview above was skipped. This will take a minute, and you'll only need to do it once. Load the protein surfaces for this section.

 

Pop3p and Pop4p share a single small edge of contact and otherwise largely splay away from each other. This accomodates Rpr2p filling in the cleft left between Pop3p and Pop4p. To best show that, Rpr1 RNA has been hidden.
The length of the contact ridge between Pop3p, and Pop4 is ~19 Šwith the surface area of contact of 233.7 Ų (6aH3) / 226.6 Ų (6agb) (Source for contact area: 'Interfaces' page of PDBePISA). You may wish to further explore the 'cementing' role for youself by toggling off-and-on Rpr2's surface using the toggles for surface at the far left. (Depending on the current state of the view, at first it may take a couple of clicks of the toggle to act. If nothing happens after several clicks, they most likely that surface is not loaded.)

 

While the bulk of Rpr2's protein-protein interaction concerns filling the cleft formed between Pop3p and Pop4p, Rpr2 also embraces a long two-stranded β-sheet, and the next few scenes explore this interaction.
Progressively fading the surface away from opaque and representing the cartoon ribbon of Pop3p with N->C rainbow coloring highlights the N- and C-termini of Pop3p come together to form an isolated, long two-stranded ribbon of antiparallel β structure. In the N->C rainbow coloring, the amino-terminus is blue and the C-terminus is red. (Bear in mind though that the observed N- and C-termini of Pop3 are short by thirteen and seven encoded residues, respectively.)
The N-terminus of Rpr2p abuts the C-terminus. The coming together of the N- and C- termini of Rpr2 directs the N-terminus of Rpr2p to twine up around Pop3p's extended β-sheet. (Note: twelve residues at the N-terminus of Rpr2 are not resolved in the structure).

 

In addition to wrapping around Pop3p's extended β-sheet, the top surface of Rpr2 is largely concerned with interacting with Rpr1 RNA. Turning back on the Rpr1 RNA for this scene starts to reveal this interaction.
Rpr2p embracing of the termini of Pop3p supports the extended β-sheet of Pop3 contacting the Rpr1 RNA.
The back of this part of this portion of Rpr2 makes contact with the pre-tRNA as well.

Rpr2p binds zinc

Just before its C-terminus, Rpr2 features a structural feature, a zinc ribbon(?), composed of two zinc ion-coordinating loops and a three-stranded beta-sheet. This single motif contacts both Pop3p and Pop4p, well above the large helix that runs along the ridge connecting the two proteins. To make it obvious, the structural feature is highlighted in red here with the zinc ion displayed as a sphere. (It is being described as a 'feature' because similar structural motifs are observed in diverse structures.)
The sheet portion, highlighted in pink, of this feature lays over Pop3p.
The two loops, with the Zn²⁺ ion sandwiched in between, interact with a small helix of Pop4. This portion of the structural feature is not colored in any special way because the zinc ion clearly indicates the feature location. (Note that here the helix of Pop4p looks to be at the N-terminus; however, the first 76 encoded residues of Pop4p are not visible in the structure with the pre-tRNA bound. The first 24 residues are visible in the structuee 6agb where pre-tRNA is not present, although there is a gap between 24 and 77.) It takes a moment, but generating the surfaces between Rpr2p (orange) and Pop4p (violet red) gives a better sense of the complementarity between the two interacting surfaces. Alternatively, now that you know where this feature is, return to the 'Rpr2p filling in the cleft' scene in the last section and toggle off-and-on 'loaded Rpr2 surface' to explore the complementarity.

 

Four cysteines directly coordinate Zn²⁺ ion binding. They are in shown in yellow wireframe here to highlight them. They are specifically residues 90, 93, 115, and 117. (Officially, the RNase P structures only list 90, 115, and 117 as binding zinc. Conservation and location of the residue corresponding to Cys93 in other higher-resolution structures [see PDB ids 1x0t and 2ki7] suggest it is part of the coordinating structure as well.)

 

 

Interaction of Pop6p and Pop7p with other components

Pop6p and Pop7p intimately contact the P3 element of the Rpr1 RNA. In fact, originally an RNA mimic of the P3 RNA element and Pop6p and Pop7p had been determined (reference goes here).
In addition to the RNA, there are additional protein-protein contacts by Pop6p and Pop7p with Pop1p, Rpp1p, and Pop5p. This can be see visualized by focusing on Pop6p and Pop7p and making any protein residues containing atoms beyond 4.5 angstoms from Pop6p or Pop7p white, as shown here. The color of residues approaching Pop6p or Pop7p are colored as the containing proteins and the cartoon loops for those residues are ehanced to make them easier to spot.
Another way to see this and get better sense of the protein packing in the region is to visualize with spacefilling van der Waals radii for all the atoms of the other proteins and making any protein residues containing atoms beyond 4.5 angstoms from Pop6p or Pop7p white, as shown here.
(You most likely want to click the 'initial-like' button below if you are trying to go back to re-run the script that turns cartoon backbone white after the spacefill one.)

 

Pop6p and Pop7p are both shared by RNase MRP and in the cryo-EM structure as well. In fact...

 

TBD  
 

 

 Schmitt
    Lab