Gallery of RIP induced Conformational Changes

One of the powerful features of RIP is that it can induce large conformational changes simply by perturbing a single sidechain. These motions are generated without the need for applying any kind of constraints at all.

For experienced MD simulators: the background fluctuations may seem rather large! This is because proteins in implicit solvent models experience no viscosity as there are no water molecules to slow down the motions.

The ATP binding loop of HSP90

HSP90 is the most common known chaperone and interacts with more proteins than any other chaperone. HSP90 functions as a elaborate machine pivoting across a dimer interface. This motion is modulated by ATP where the N-terminal domain binds ATP through a huge loop. This loop is known to undergo large conformational changes as it has been crystallized in many different conformations. Applying RIP to a residue on the ATP-binding loop, we can induce the loop to move several Åstroms of motion. (flexibility analysis of N-terminal domain of HSP90).

The Ligand-Binding Loop of Triosephosphate Isomerase

Triosephosphate Isomerase (TIM) is a classic β-barrel domain. This fold provides a sturdy scaffold for many catalytic reactions as the middle of the barrel forms a great catalytic site where sidechains have easy access to the ligand. As well, TIM has a very well- characterized loop that undergoes 6 to 7 Ångstroms of motion to bind the ligand, which is crucial for catalysis. Here we induce a large conformational change in the loop by applying RIP to a neighboring glutamate. (flexibility analysis of the open conformation of TIM).

The Zinc-Finger Mimic Falling Apart

This classic artificially-designed protein, designed in the laboratory of Stephen Mayo, folds into the classic zinc finger motif without the need for any zinc ions at all! It is an extremely small protein of 28 amino acids. If RIP is applied to a key residue in the hydrophobic core, the protein falls apart. (flexibility analysis of zinc-finger mimic).

Allosteric Helix-12 of the Ligand-Binding Domain of the Estrogen Receptor

The Estrogen Receptor is a complex machine that triggers large-scale nuclear activity upon binding of Estrogen in the ligand-binding domain. Binding of ligand involves the closing of Helix-12 over the ligand. In vivo, it appears that HSP90 is needed to keep Helix-12 from closing onto the ligand-binding site when the site is empty. In this simulation starting from the closed conformation of the protein, applying RIP to a highly conserved Trp knocks out the entire Helix-12, which is otherwise well-bound to the protein. (flexibility analysis of the ligand-binding domain of ER).

The Flexible Helices of the PDZ domain of Protein Tyrosine Phosphotase

The PDZ domain is a common structural domain. It possesses several well-defined protein-protein interaction surfaces and a canonical C-terminus/peptide binding cleft along the groove of the major α-helix. Many scaffold proteins consists of a string of PDZ domains that make use of these protein interaction sites to assemble related proteins together. Once thought to be passive, it is now known that certain PDZ domains undergo allostery, such as the second domain of Protein Tyrosine Phosphatase. NMR studies suggests that both helices in this protein are mobile until ligand binds. Presumably, rigidification of these α-helices activates the protein-protein interaction surfaces defined by the α-helices. In this example, a RIP perturbation near the peptide-binding site kicks out the smaller α-helix, demonstrating the extreme flexibility of this α-helix. (flexibility analysis of 2nd PDZ domain of PTP).

The Exaggeration of Electrostatics

One of the down sides of using implicit solvent force-fields is that electrostatics on the surface of the protein are over-exaggerated. In this example, a lysine along an α-helix forms a strong temporary salt-bridge with a glutamic acid that shakes out the whole helix.