Not just for cleaving barns anymore!!
Barnase is a 110-residue extraceullular protein found in bacillus amyloliquefaciens. It is a ribonuclease whose potentially lethal functions within the cell are inhibited by barstar, a 90-residue polypeptide. Barstar binds tightly to barnase through salt bridges and hydrogen bonds, inhibiting its RNase activity. Once bound to barnase, barstar sterically blocks the active site of barnase with an alpha helix and the loop segment connecting it to the adjacent helix.
Formation of the barstar-barnase complex is dependent on pH as evidenced by the presence of more than 35 water molecules within 4.5 angstroms of both protein molecules. The structure of barnase experiences only minimal changes when bound to barstar. Again, barstar functions by blocking the active site rather than affecting conformational changes. The changes that do occur in barnase are limited to the region localized around the barnase-barstar complex. When bound, however, the two lobes of barstar expand slightly to accomodate barnase.
To gain a better understanding of barstar and how it functions, we will take you through a tutorial of barstar using pictorial representations.
The first view shows the main back bone of the structure. To view, click the button blow:
The alpha helixes can be seen easily in this view. The beta sheets can be visualized better if the molecule is rotated. By clicking on the button below you will be able to see all the main chain atoms color coded by the different atoms.
From this view the alpha helixes and the beta sheets can still be seen by rotating the molecule. Now we will give you the best way of viewing the secondary structure of barstar. To look at the protein with the structures color coded in cartoon form, click below.
Looking at the secondary structure, you can see clearly that there is also a supersecondary structure to this protein. There is a motif of beta-alpha-alpha-beta. By simply rotating with a left mouse click, the supersecondary structure can be seen on both monomers. To get a closer look, hold shift and scroll down with the mouse. Click below to reset the molecule.
Next we will look at the acidic and basic sites of barstar. The basic sites will be shown in purple and the acidic sites are shown in yellow.
Next we will look at the charged areas on the protein. Clicking on the button below will display the charged atoms in the protein. All the charged side chains are pretty well spread out over the exterior of the entire molecule with some charged side chain localized in between the two segments of barstar. When bound, unfavorable electrostatic interactions are stabilized through the trapping and burial of solvent molecules.
Click on the button below to reset the molecule.
Next, we'll examine the degree of hydrophobicity. To do this, click on the button below. This will display the hydrophobic areas of the protein in bloody cool colors. As you can see, the hydrophobic residues are buried in the protein. Again click below to reset the protein.
To get a better understanding of the hydrophobic effect in barstar, click the button below to see the hydrophobic regions in wireframe.
Click the "Original Settings" button below to reset the molecule.
To see where the solvent accessable sights are on the Barstar click below. It may be a little hard to see, but just scroll around to see the sovlent dots!
Click below to reset the molecule!
Barnase and barstar interact electrostatically through 14 hydrogen bonds between residues:
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
The two phosphate binding sites in barnase are occupied in the barnase-barstar complex by Asp39 and Gly43 of barstar. Now we will get a better look at the barstar and barnase interaction click on the button below to see basrstar and barnase together. From this you can see that ther in fact two barstar proteins for every one barnase. Clicking on the button labeled See hydrogen bonds and secondary structure, and you will get a better look!
You can clearly see that the interaction between the barstar and barnase is through a alpha helix and the barstar and a beta sheet on the barnase.
To get a even better look at the active sight click below!!
The picture below shows the minimized structure of barstar and Charmm energy from the Quanta/Charmm program
The picture below is the Ramachandran plot of the Phi/Psi angle, showing which angles allow for the necessary conformational space. The purple areas represent the permitted right-hand alpha helix Phi/Psi angles. The yellow and red regions represent the parallel and anti-parallel beta sheets.
The picture below shown the barnase in space filling form, and the barstars in ribbon form. This shows how the barstar ligands come in and sterically block the active site of barnase, thus inhibiting it.
That's all, folks!!!