User:Loganne Wertz/Sandbox1
From Proteopedia
Contents |
Zymogen Activation
In addition to a self-cleavage mechanism, Caspase-6 zymogen can be activated getting cleaved by Caspase-3, as well as other enzymes. The mechanism of activation by clevage is highly conserved across the caspase family; Self-processing is uniquely recognized as the primary mechanism for Caspase-6 activation, where clevage must occur at two sites for complete activation, specifically the pro-domain and the intersubunit linker. These cleavages are both sequence specific and ordered. First, pro-domain must be cleaved. (Some residues of the pro-domain are not visible in the crystallized structure) Then cleavage of the intersubunit linker occurs, cleaving both DVVD179 and TEVD193. To some extent the pro-domain inhibits Caspase-6's ability to cleave the intersubunit loop and self-activate; It has been proposed that this sequence of cleavage is due to the pro-domain being more readily available to enter the active site. The result of the TETD23 cleavage site priority is that the prodomain acts as a “suicide protector”, which protects the TEVD193 cleavage site from self-cleavage[3]. This protection is necessary when there are low levels of inactive proteins, which must be preserved, in the tissue. The intramolecular cleavage of TETD23 and DVVD179 or TEVD193 are essential for the initiation caspase-6 activation without other caspases present. After both cleavages occur, the processed Caspase-6 can be found in solution as a dimer of dimers.
Active State
In order to function as an endoprotease, Caspase-6 binds a , which can include neuronal proteins and tubulins [1], in its active site.
Zinc Inhibition
Caspase-6 function is primarily inhibited by the binding of a ion[2], which binds to an instead of the . This allosteric site is located on the outside of the protein and is distal to the active site. The zinc ion is bound to , Lys-36, Glu-244, and His-287. Once the ion is bound to the protein, it is then stabilized by a found in the cytoplasm. The binding of zinc at the exosite is suggested to cause a conformational change in the protein from an to an that misaligns catalytic residues and inhibits activity of the enzyme. It has been proposed that helices of the active dimer must rotate or move in some other way to provide these ideal interactions with zinc. This subtle shift is most likely the cause for allosteric inhibition. As the helices move to bind zinc, the amino acids of the active site become misaligned. The altered positions of the amino acids no longer provide ideal interactions for incoming substrates. After zinc binds, no new substrates enter the active site. Thus, Caspase-6 is effectively inhibited. The residues in the active site no longer provide ideal interactions with the substrate and therefore, substrate does not bind. Zinc binding to the exosite is tightly regulated as it inhibits Caspase-6's critical role in initiation of apoptosis.
Phosphorylation
The function of Caspase-6 can be inhibited by phosphorylation of Ser-257. The exact mechanism of this reaction remains unidentified at the time of publication, but proceeds when ARK5 kinase is present. This modification can occur before and after zymogen activation. The phosphoryl group inhibits Caspase-6 through steric interference. When Ser-257 is phosphorylated, the amino acid residue interacts with , causing a shift in the helices of Caspase-6. This is shown in the , whose mutation mimics phosphorylation. [1] The shift misaligns and disrupts residues found in the active site. This conformational difference prevents the intersubunit linker from entering during zymogen activation and the self-cleaved active dimer cannot be formed. Additionally, no new substrate is able to enter the active site.
