Shape shifting: The multiple conformational substates of the PTEN N‐terminal PIP₂‐binding domain

Abstract

The Phosphatase and TENsin homolog deleted on chromosome 10 (PTEN) is a chief regulator of a variety of cellular processes including cell proliferation, migration, growth, and death. It is also a major tumor suppressor gene that is frequently mutated or lost under cancerous conditions. PTEN encodes a dual‐specificity (lipid and protein) phosphatase that negatively regulates the PI3K/AKT/mTOR signaling pathway where the PIP2‐binding domain (PBD) regulates the lipid phosphatase function. Unfortunately, despite two decades of research, a full‐length structure of PTEN remains elusive, leaving open questions regarding PTEN's disordered regions that mediate protein stability, post‐translational modifications, protein–protein interactions, while also hindering the design of small molecules that can regulate PTEN's function. Here, we utilized a combination of crosslinking mass spectrometry, in silico predicted structural modeling (including AlphaFold2), molecular docking, molecular dynamics simulations, and residue interaction network modeling to obtain structural details and molecular insight into the behavior of the PBD of PTEN. Our study shows that the PBD exists in multiple conformations which suggests its ability to regulate PTEN's variety of functions. Studying how these specific conformational substates contribute to PTEN function is imperative to defining its function in disease pathogenesis, and to delineate ways to modulate its tumor suppressor activity.

Structural and Dynamic Effects of PTEN C-Terminal Tail Phosphorylation

“…A modest increase in α-helical formation is seen in the PBD of PTEN 4A, which persists throughout entire MD simulation yet increases from 9% in the WT PTEN system to 32% in the PTEN 4A system (Table S5 and Figure A,C), which suggests that CTT mutation at positions S380/T382/T383/S385 to alanine destabilizes the overall structure of PTEN. Overall, our in silico MD simulations show that for WT PTEN, the N-terminal PBD exists in an α-helical conformation, which agrees with experimental results , and is also consistent with our integrative structural modeling and membrane MD simulation studies . However, our results reveal that phosphorylated CTT increases the presumed α-helical character in the N-terminal PBD and IDR regions of PTEN, which is in contrast to recent NMR studies where they demonstrate a phosphorylated CTT peptide diminishes the α-helical character of the N-terminus.…”

Comparative Protein Structural Network Analysis Reveals C-Terminal Tail Phosphorylation Structural Communication Fingerprint in PTEN-Associated Mutations in Autism and Cancer

“…Interestingly, though M134R and R173C have a global metapath that extends from the phosphatase domain across the inter-domain and into the β-sandwich of the C2 domain, the P-M134R and P-R173C systems have less recurrent links and a more clustered global metapath, concentrated in their core (inter-domain interface). This supports our previous results where cancer-associated mutations demonstrated a long-range communication pathway within the inter-domain region of the protein and underscores the critical functional roles of the Met134 (M134) and the highly conserved Arg173 (R173) positions in allosteric communication. , M134 mutations result in compromised lipid phosphatase activity , and are associated with breast cancer and Bannayan–Riley Ruvalcaba syndrome, , supporting its functional importance. Moreover, R173, which forms a rich hydrogen bond network with the inter-domain region, is among the eight most frequently mutated PTEN residues in PHTS-cancer …”

“…For each structure production run, 1000 ns each (1 μs) of unrestrained NPT was performed at 303 K, where bond lengths were constrained using the linear constraint solver (LINCS) algorithm (Table S1, see Supporting Information). Our simulations closely followed the same protocol as in our previous works. ,, Coordinates were saved every 20 ps, leading to 50,000 configurations for each system. Trajectory analyses were conducted using GROMACS utilities, VMD or PyMOL…”

“…Our simulations closely followed the same protocol as in our previous works. 8 , 9 , 26 Coordinates were saved every 20 ps, leading to 50,000 configurations for each system. Trajectory analyses were conducted using GROMACS utilities, VMD 27 or PyMOL.…”

“…This supports our previous results where cancer-associated mutations demonstrated a long-range communication pathway within the inter-domain region of the protein and underscores the critical functional roles of the Met134 (M134) and the highly conserved Arg173 (R173) positions in allosteric communication. 6,26 M134 mutations result in compromised lipid phosphatase activity 49,50 and are associated with breast cancer 49 and Bannayan−Riley Ruvalcaba syndrome, 49,51 supporting its functional importance. Moreover, R173, which forms a rich hydrogen bond network with the inter-domain region, is among the eight most frequently mutated PTEN residues in PHTS-cancer.…”