The is a human poly (ADP-ribose) glycohydrolase.It is the major enzyme responsible for the catabolism of poly (ADP-ribose), a reversible covalent-modifier of chromosomal proteins. The protein is found in many tissues and may be subject to proteolysis generating smaller, active products.
This protein is only present when the DNA is damaged. It influences the damaged chromatin through a derepression of a gene promoter. Consequently this protein is quite interesting for biotechnological applications. Indeed, knowing the different pathways and protein interactions leading to DNA damage repair is a meaningful goal in research especially in new cancer therapies.
Structure
Structural highlights
This protein has four principal domains on a : an A-domain, a catalytic domain and two substrate binding domains[1].
The first 456 amino acids of the peptide chain form the . Then, from the 610th to the 795th amino acids the can be found. This catalytic domain can bind to other proteins with (the 726th and 727th amino acids). Next, is located from the 869th to the 874th amino acid. [2]
As such, most of the amino acids form the A-domain and the catalytic domain and only a few amino acids (8 a.a) make links with other proteins. Moreover, the ligand 7JB can bind the protein on the . These amino acids are located on a helix and on a beta sheet. [3]. There is the amino acid distribution.
Secondary Structure
This protein is 37% and 13% , distributed . Indeed, 6HMM has 25 helices on 198 residues and 23 strands on 74 residues. It also has a few 3/10 helices. [4]
Concerning the torsion angles, the backbone angles and the sidechain angles can be differentiated. Indeed, no residue in the does not respect the Ramachandran's angle, whereas only 2% of the residues on the sidechain are Ramachandran outliers due to having a non-rotameric form. [5]
Tertiary Structure
The protein PARG folds into an ADP-ribose-binding macro domain with an N-terminal extension. It also consists of a diphosphate-binding loop on one side of an ADP-ribose binding cavity. On the other side there are several amino acids matching to the specific PARG signature sequence.
In the macro domain fold, a loop is inserted to welcome the Glu115 side chain protecting the active site of the PARG protein. This loop gives PARG the ability to hydrolyze PAR.
Concerning the ligand pairing with the PARG protein only a small difference can be observed for the amino acids Val226 and Phe227 [1].
Quaternary Structure
The Poly(ADP-ribose)glycohydrolase can interact with either PCNA or NUDT5, this gives various possible functions to the protein. When this protein binds with NUDT5 it can remodel chromatin for example [6].
Function
The protein is a complex composed of the poly (ADP-ribose) glycohydrolase (PARG) and the anthraquinone PDD00013907. The post-translational modifications of the PAR protein (poly ADP-ribose) are important for DNA stability.
PDD00013907 is, as already stated, an anthraquinone which is a polycyclic aromatic hydrocarbon usually used in biopesticides as a pest repellant. Here it is considered as a free (of identification number on PDB: 7JB) that can bind to the creating the.
Post-translational modifications and anthraquinone
There are several possible post-translational modifications to stabilize DNA. Most commonly there would be phosphorylation, acetylation or methylation [2]. Another post-translational modification concerning the 6HMM protein is made on the poly(ADP-ribose) protein (PAR). PAR is composed of a repetition of ADP-ribose units linked through glycosidic ribose-ribose bonds [3]. This allows the repair of single-strand breaks on DNA [4]. PARG, a constituent of the 6HMM protein, will degrade PAR to allow the poly (ADP-ribose) polymerase (PARP) to free itself from the damaged site, that is now repaired, and completes as such reparation [5].
The anthraquinone PDD00013907 is a weakly active and cytotoxic anthraquinone 8a acting as a free ligand binding in the ADP-ribose binding site of the PARG. This PDD00013907 should lead to the inhibition of PARG, which is of interest in the search of novel cancer therapies [5].
The mechanism
As said previously, poly(ADP-ribosylation) is an important post-translational modification for DNA repair. The mechanism behind this repair relies on several factors. At first, the Poly (ADP-ribose) polymerase (PARP), more specifically the subtype PARP1, will recognize and will bind to the single-stranded break on the DNA. It will then autophosphorylate due to NAD+ and form PAR chains. These will recruit other repair proteins to the site.
The role of PARG is the hydrolyzation of the specific ribose-ribose bonds present in PAR which leads to its degradation and as such the reparation cycle will be finished[4]. This degradation is important because without PARG the repair cycle cannot be completed [6] and may lead to cell death. This is partially due to the still present PARP on the previously damaged site maintained by the non-degraded PAR [7].
Diseases and Relevance
Due to the function of the protein poly (ADP-ribose) glycohydrolase PARG to be part of post-translational processes of DNA damage repair it could be used for new treatments in cancer therapy or for ther diseases. In cancer cells the rate of DNA damage is most probably higher than in normal cells. This could result from the considerably raised stress levels. A deficiency of PARG results in the cessing of the cell cycle and the following cell death[4][5]. Consequently, the inhibition of PARG might be a solution on how to destroy tumor cells, for example.
For the PAR protein, there has already been a lot of research concerning novel treatments, but not for PARG. As there are no close homologues of PARG[4], this protein provides a potential target in drug discovery.
As the protein in complex with the anthraquinone does not work properly anymore [7], the described complex shows how an inhibited PARG might act in the cell. The goal of searched therapeutics is therefore to find a way to get the protein PARG into a complex that is acting similar to the 6HMM complex. For now, the research for anthraquinone as inhibitor has stopped as it is cytotoxic for the cell[5].
