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Parkinson’s disease is one of the most common neurodegenerative diseases worldwide. In comparison, Parkinson’s is ranked just below Alzheimer’s disease, with a 2% lifetime risk. This disorder causes neurons in the body to change their structure dramatically and lose function. These changes trigger symptoms that affect a person’s motor skills, which can include tremors, rigid muscles, slowed movement, change of speech, and weakened balance. While Parkinson’s disease is known for its prevalence with motor skills, it can also affect judgment, decision-making, and memory. Currently there are treatments to manage this disease, but a lot of progress needs to be made to find a cure (Schapira).
Leucine rich repeat kinase 2 (LRRK2) is a key protein that can be useful in the treatment of Parkinson’s disease, but currently researchers do not fully comprehend how it specifically correlates to treatment. However, the majority of cases have a similar mutation, G2019S , in the kinase domain of LRRK2, which has proven to increase kinase activity. The idea is to detect kinase inhibitors that can reduce the activity of this mutation in hopes that it will lead towards a better treatment. A kinase is an enzyme that activates phosphate groups to transition to a substrate from high-energy molecules. In order to find more information about the kinase crystal structure of LRRK2, tyrosine kinase 2 () was used as a model because it is 74% similar in ATP-binding residues to LRRK2 (Galatsis).
Although, the kinase domain does seem the most relevant to the treatment of Parkinson’s disease, the ROC domain of LRRK2 has proven to regulate kinase activity as well. It does this by yielding GDP from GTP through alternations in its conformation in a GTP-bound and GDP-bound cycle. The GDP-Mg2+ ligand in the ROC domain plays an important role in the complex by stabilizing GDP (Deng).
Overall Structure
The Overall Structure of LRRK2 is shown in the window on the right and may be returned to by clicking .
The two chains, or units, that make up LRRK2 are essentially each TYK2 proteins, meaning LRRK2 is a dimer of TYK2. The Overall Structure of TYK2 is shown . The one major structural difference is that LRRK2 includes 2 Mg+ ions while Tyk2 does not.
- Building the LRRK2 Protein
There are 12 beta sheets that form the backbone of the protein. to add them. 6 sheets belong to Chain #1 and 6 sheets belong to Chain #2 . to differentiate each chain. They are organized into 4 groups of 3 sheets, where each group is adjacent to a group belonging to the opposite chain. This ordering increases the total protein stability by interweaving the two chains, but still allows for hinged movement.
LRRK2 additionally contains 10 alpha helices. The helices surround the beta sheets like a turtle’s shell, covering the beta sheets with the exception of one large side (belly) and the centers on the two long ends (head and tail). to add the alpha helices . to differentiate each chain.
Now we to complete the protein chain.
Now all that is left is to that are featured in the overall structure. There are 2 Guanosine Diphosphate compounds shown in ball and stick representation, along with 2 Magnesium ions, represented as spheres.
- Location of Hinge Structures
Each subunit of LRRK2 contains a single strand that connects the alpha helices to the beta sheets. These may act as hinges during binding interactions.
- Polar and Non-polar Groups
For LRRK2, when looking at the , you see that most residues are polar there are very few non-polar .
Conversely, the consist almost entirely of non-polar groups.
Binding Interactions
LRRK2 contains binding sites for 6-[(2,5-dimethoxyphenyl)sulfanyl]-3-(1-methyl-1H-pyrazol-4-yl)[1,2,4]triazolo[4,3-b]pyridazine which we will refer to by its PDB code, 2YK.
The residues involved in binding interactions are listed below:
- LEU 903A
- VAL 911A
- GLY 1040A
- GLU 979A
- VAL 981A
These residues are highlighted in green and the ligand 2YK is shown in blue.
The scene below provides an alternative view of the binding interactions with the residues labelled in red with their corresponding number and the ligand, 2YK, labelled in blue.
Additional Features
LRRK2 is a very large protein comprised of multiple domains. The most well-known LRRK2 mutation occurs in the kinase domain and plays a role in Parkinson’s disease by increasing its kinase activity.
Researchers have since discovered that the ROC domain also regulates LRRK2’s kinase activity by functioning as a GTPase which switches between an active GTP- and inactive GDP-bound conformation[1]. The for both monomers is an Mg2+ binding pocket located on the surface of the ROC dimer, each shown with a bound GDP. Although the mechanism is unclear, LRRK2 is believed to exhibit intramolecular regulation: the activity of LRRK2’s kinase domain is stimulated by the active GTP-bound state of its own ROC domain.
Two residues whose mutations are associated with Parkinson’s disease, R1441 and I1371, are the key players of LRRK2’s GTPase-regulated kinase activity, because they work together to stabilize the ROC dimer [2]. Specifically, the guanidium group of R1441 exhibits essential hydrogen bonding with the backbone carbonyl oxygen of residue F1401 and the hydroxyl group of the other monomer’s nearby alpha helix. Additionally, the guanine group, W1434’s six membered-ring, and the side chains of F1401 and P1406 create a hydrophobic “zipper” via stacking interactions. The I1371 residue is located within a hydrophobic pocket and provides optimal Van der Waals interaction with T1404’s methyl group.
Due to the intricate hydrogen bonding, hydrophobic contacts, and other important interactions contributed by R1441 and I1371, mutations of either residue can easily destabilize the ROC dimer and alter its function. Research demonstrates that these mutations disrupt the hydrolysis of GTP to GDP, prolonging the active GTP-bound state[3]. Ultimately, this reduction in GTPase function of the mutated ROC dimer increases LRRK2’s kinase activity, a classic marker of Parkinson’s disease.
Quiz Question 1
The molecule shown is TYK2. Since the structure of TYK2 is almost identical to the structure of the kinase region in LRRK2, we will use it to simplify the concept of the .
The is the only segment that connects the (mainly) to the (mainly) . Various intramolecular interactions between the upper and lower segments help the protein maintain its tertiary structure. The strength of the interactions determines the rigidity of the hinge. If we were to add several cysteine molecules that were able to form disulfide bonds between the upper and lower segments without disrupting the active site, how would the kinase be affected? In your description, include whether the mutations would primarily affect Km or Vmax, how they would affect these kinetic parameters, and give a brief explanation of why.
Quiz Question 2
For our second question I am going to present a green scene with the binding pocket shown and representation of the ligand shown. The green scene will show the details of the binding pocket including the secondary structures and the residues which make them up. I will then ask the student to think about the scene and give three ways in which they could modify the ligand which would make it a better competitive inhibitor.
See Also
Credits
Introduction - Megan Greiner
Overall Structure - Nick Barberio
Drug Binding Site - John Vetrano
Additional Features - Nicole Garvin
Quiz Question 1 - Charit Tippareddy
Quiz Question 2 - Peter Kelly
References
- ↑ Guo L, Gandhi PN, Wang W, Petersen RB, Wilson-Delfosse AL, Chen SG. The Parkinson's disease-associated protein, leucine-rich repeat kinase 2 (LRRK2), is an authentic GTPase that stimulates kinase activity. Exp Cell Res. 2007 Oct 1;313(16):3658-70. Epub 2007 Jul 19. PMID:17706965 doi:http://dx.doi.org/10.1016/j.yexcr.2007.07.007
- ↑ Deng J, Lewis PA, Greggio E, Sluch E, Beilina A, Cookson MR. Structure of the ROC domain from the Parkinson's disease-associated leucine-rich repeat kinase 2 reveals a dimeric GTPase. Proc Natl Acad Sci U S A. 2008 Feb 5;105(5):1499-504. Epub 2008 Jan 29. PMID:18230735
- ↑ Li X, Tan YC, Poulose S, Olanow CW, Huang XY, Yue Z. Leucine-rich repeat kinase 2 (LRRK2)/PARK8 possesses GTPase activity that is altered in familial Parkinson's disease R1441C/G mutants. J Neurochem. 2007 Oct;103(1):238-47. Epub 2007 Jul 10. PMID:17623048 doi:http://dx.doi.org/10.1111/j.1471-4159.2007.04743.x
