Үбіквитинды кесу ферменттері: жіктемесі мен қызметі

Deubiquitinating enzymes (DUBs), also known as deubiquitinating peptidases, deubiquitinating isopeptidases, deubiquitinases, ubiquitin proteases, ubiquitin hydrolases, or ubiquitin isopeptidases, are a large group of proteases that cleave ubiquitin from proteins. DUBs can reverse these effects by cleaving the peptide or isopeptide bond between ubiquitin and its substrate protein. In humans there are nearly 100 DUB genes, which can be classified into two main classes: cysteine proteases and metalloproteases. The cysteine proteases comprise ubiquitin specific proteases (USPs), ubiquitin C terminal hydrolases (UCHs), Machado Josephin domain proteases (MJDs) and ovarian tumour proteases (OTU). The metalloprotease group contains only the Jab1/Mov34/Mpr1 Pad1 N terminal+ (MPN+) (JAMM) domain proteases.

The Jab1/Mov34/Mpr1 Pad1 N terminal+ (MPN+) (JAMM) domain superfamily proteins bind zinc and hence are metalloproteases. Proteins are affected by these modifications in a number of ways: they regulate the degradation of proteins via the proteasome and lysosome; coordinate the cellular localisation of proteins; activate and inactivate proteins; and modulate protein protein interactions. DUBs cleave the ubiquitin from these proteins, producing active single units of ubiquitin. Free polyubiquitin chains are cleaved by DUBs to produce monoubiquitin. The chains may be produced by the E1 E2 E3 machinery in the cell free from any substrate protein. Another source of free polyubiquitin is the product of ubiquitin substrate cleavage. If DUBs cleave the base of the polyubiquitin chain that is attached to a protein, the whole chain will become free and needs to be recycled by DUBs.

The catalytic domain of DUBs is what classifies them into particular groups; USPs, OTUs, MJDs, UCHs and MPN+/JAMMs. The first 4 groups are cysteine proteases, whereas the latter is a zinc metalloprotease. The cysteine protease DUBs are papain like and thus have a similar mechanism of action. They use either catalytic dyads or triads (either two or three amino acids) to catalyse the hydrolysis of the amide bonds between ubiquitin and the substrate. The active site residues that contribute to the catalytic activity of the cysteine protease DUBs are cysteine (dyad/triad), histidine (dyad/triad) and aspartate or asparagine (triad only). The histidine is polarised by the aspartate or asparagine in catalytic triads or by other ways in dyads. This polarised residue lowers the pKa of the cysteine, allowing it to perform a nucleophilic attack on the isopeptide bond between the ubiquitin C terminus and the substrate lysine. Metalloproteases coordinate zinc ions with histidine, aspartate and serine residues, which activate water molecules and allows them to attack the isopeptide bond.

Ubiquitin like (UBL) domains have a similar structure (fold) to ubiquitin, except they lack the terminal glycine residues. 18 USPs are proposed to have UBL domains. Only 2 other DUBs have UBLs outside the USP group: OTU1 and VCPIP1. USP4, USP7, USP11, USP15, USP32, USP40 and USP47 have multiple UBL domains. Sometimes the UBL domains are in tandem, such as in USP7 where 5 tandem C terminal UBL domains are present. USP4, USP6, USP11, USP15, USP19, USP31, USP32 and USP43 have UBL domains inserted into the catalytic domain. The functions of UBL domains are different between USPs, but commonly they regulate USP catalytic activity. They can coordinate localisation at the proteasome (USP14); negatively regulate USPs by competing for the catalytic site of the USP (USP4), and induce conformational changes to increase catalytic activity (USP7). Like other UBL domains, the structure of USP UBL domains show a β grasp fold.

Single or multiple tandem DUSP domains of approximately 120 residues are found in six USPs. The function of the DUSP domain is currently unknown but it may play a role in protein protein interaction, in particular to DUBs substrate recognition. This is predicted because of the hydrophobic cleft present in the DUSP domain of USP15 and that some protein interactions with DUSP containing USPs do not occur without these domains. The DUSP domain displays a novel tripod like fold comprising three helices and an anti parallel beta sheet made of three strands. This fold resembles the legs (helices) and seat (beta sheet) of the tripod. Within most DUSP domains in USPs there is a conserved sequence of amino acids known as the PGPI motif. This is a sequence of four amino acids; proline, glycine, proline and isoleucine, which packs against the three helix bundle and is highly ordered.

The full extent of the role of DUBs in diseases remains to be elucidated. Their involvement in disease is predicted due to known roles in physiological processes that are involved in disease states; including cancer and neurological disorders. The enzyme USP28 is over expressed in different types of cancer such as colon or lung. In addition, USP28 deubiquitinates and stabilizes important oncogenes such as c Myc, Notch1, c jun or ΔNp63. In squamous tumors, USP28 regulates the resistance to chemotherapy regulating DNA repair via ΔNp63 Fanconia anemia pathway axis. The deubiquitinating enzymes UCH L3 and YUH1 are able to hydrolyse mutant ubiquitin UBB+1 despite the fact that the glycine at position 76 is mutated. UCH L1 levels are high in various types of malignancies (cancer).

DUBs play an active role in modulating the cell cycle. Ubiquitin specific processing protease (USP) is a family of deubiquitinating enzymes that play a crucial role in cell cycle regulation. Two such enzymes include USP17 and USP44. USP17 regulates pathways responsible for progressing cells through the cell cycle. Its targets include regulators of Ras, CDK2, and Cyclin A. USP44 plays an important role in anaphase initiation. New research into the mitotic checkpoint has revealed a novel role for USP44 in regulating cell cycle progression. USP17 acts to deubiquitinate K63 ubiquitin domains on RCE1. Thus, regulation of Ras through USP17 acts as another point in Ras regulation.

Cyclin dependent kinases (CDKs) are a family of enzymes that phosphorylate serine and threonine residues to drive the cell through the cell cycle. Activation of CDK2 is critical for the G1 S transition. For CDK2 to be activated, cyclin A must bind to the cyclin dependent kinase complex (CDKC). Cell division cycle 25A (CDC25A) is a phosphatase that removes an inhibitory phosphate group from CDK2. While ubiquitination would mark CDC25A for degradation, thus blocking progression to S phase, USP17 deubiquitinates CDC25A. This compaction of the DNA downregulates CDKN1A transcription. USP17 deubiquitinates SETD8, thus reducing its propensity for degradation and increasing its intracellular stability. Such a mechanism is crucial, as errors in chromosomal separation have been implicated in cancer, birth defects, and antibiotic resistance in pathogens. One of the core regulator proteins is the anaphase promoting complex (APC/C). APC/C ubiquitinates securin. The resulting destruction of securing release separase, A protein called hMAD2 can form an inactive trimer with APC and CDC20, forming the hMAD2 CDC APC complex. It is important to note that ubiquitination of CDC20 does not serve to mark it for degradation, but rather promote dissociation of hMAD2 from the hMAD2 CDC APC complex. USP44, a ubiquitin specific processing protease, can stabilize the inactive hMAD2 CDC APC complex by counteracting UbcH10 ubiquitination. This blocks hMAD2 dissociation and allows for proper regulation of APC/C, keeping it inactive until proper attachment of the mitotic spindle. Upon proper attachment, switch like behavior allows for the activation of APC/C. Deubiquitinating enzymes play an integral role in maintaining p53’s function. In healthy cells, p53 activates the E3 ubiquitin ligase MDM2 which in turn ubiquitinates p53. This creates a negative feedback loop, whereby the degradation of p53 allows for cells to flow through the cell cycle. Upon DNA damage, Ubiquitin specific processing protease 7 (USP7) stabilizes p53 by cleaving ubiquitin. For USP7 to deubiquitinate p53, it must localize to the nucleus. However, no nuclear localization sequence (NLS) has been found. Despite no known NLS, one study showed that, upon deletion of USP7’s N terminus, no nuclear localization occurred. or promote cell death, depending on the severity of the DNA damage. See schematic of the role of USP7 in the p53 dependent pathway. or promote cell death, depending on the severity of the DNA damage. See schematic of the role of USP7 in the p53 dependent pathway.