Phosphorylation by PINK1 Releases the UBL Domain and Initializes the Conformational Opening of the E3 Ubiquitin Ligase Parkin
Figure 2
phospho-Ser65 triggers cleft widening in the N-terminal region.
A) Center view of a newly defined pocket in Parkin with Ser65 positioned towards the middle. The cleft is formed between the flexible linker region [cleft wall 1: Arg97, Ser110, Val105, Val111, Leu112, Asp115 Ser116, Val117, and Gly118] and the UBL domain [cleft wall 2: Met1, Ile2, Val3, Phe4, Ser19, Leu61, Asp62, Gln63, Gln64, Ile66, and Val67]. Parkin is rendered in solvent accessible surface colored by atom type (nitrogen-blue, oxygen-red, carbon-cyan). Yellow double-sided arrow indicates cleft regions that were used for center-of-mass calculations: CoM1 (Ser110, Val111, Asp115) to CoM2 (Met1, Ile2, Val3, Phe4, Gln63). The initial cleft width is given for time equal zero. B) The ribbons diagram for Parkin is shown for comparison with same orientation. Relevant domain labels are given. C) Parkin pSer65 is shown after 20 ns of unbiased MDS. The yellow arrow indicates the increased cleft distance. D) Superposition of Parkin structures after 100 ns MDS. Shown are structures for Ser65 and pSer65 as well as for S65A, S65D and S65E variants. Arrows indicate the cleft distances of Ser65 and pSer65. E) Plot for CoM1 to CoM2 distance over time from MDS. Graph shows a relatively closed and stable cleft for unmodified Ser65 (black) of about 8 Å in distance, while pSer65 (green) shows a much more wider cleft from the start, ranging from 11–14 Å over time. Phospho-mimic mutants S65D (magenta) or S65E (blue) as well as the phospho-dead mutant S65A (red) show a strong increase in cleft size over time reaching distance observed with pSer65. F) Solvent-Accessible-Surface-Area (SASA-Å2) within the pocket enclosing Ser65 measured in Å2 units. While Ser65 maintain a relatively stable SASA, values for pSer65 strongly increase over time. An increase in SASA over time is also observed for the substitutions S65D and S65E as well as for S65A after an initial decrease. G) Shown are numbers of H2O molecules within the cavity surrounding Ser65 during MDS. While Ser65 constantly maintains twelve H2O molecules, pSer65, S65D, S65E, and S65A show a greater solvation of the cavity, consistent with an increased cleft distance and improved SASA.