Crystal structure and molecular dynamics of human POLDIP2, a multifaceted adaptor protein in metabolism and genome stability

Abstract

Polymerase δ-interacting protein 2 (POLDIP2, PDIP38) is a multifaceted, "moonlighting" protein, involved in binding protein partners from many different cellular processes, including mitochondrial metabolism and DNA replication and repair. How POLDIP2 interacts with many different proteins is unknown. Towards this goal, we present the crystal structure of POLDIP2 to 2.8 Å, which exhibited a compact two-domain β-strand-rich globular structure, confirmed by circular dichroism and small angle X-ray scattering approaches.POLDIP2 comprised canonical DUF525 and YccV domains, but with a conserved domain linker packed tightly, resulting in an "extended" YccV module.A central channel was observed, which we hypothesize could influence structural changes potentially mediated by redox conditions, following observation of a modified cysteine residue in the channel. Unstructured regions were rebuilt by ab initio modelling to generate a model of full-length POLDIP2. Molecular dynamics simulations revealed a highly dynamic N-terminal region tethered to the YccV-domain by an extended linker, potentially facilitating interactions with distal binding partners. Models of POLDIP2 complexed with two of its partners, PrimPol and PCNA, indicated that dynamic flexibility of the POLDIP2 N-terminus and loop regions likely mediate protein interactions.

New insights in the molecular regulation of the NADPH oxidase 2 activity: Negative modulation by Poldip2

“…4-7%), a higher percentage of β-sheets (15-23%), and turns (15-22%), and a high amount of unordered secondary structure (> 56%). This SR-CD analysis is consistent with the high β-strand content CD spectra obtained for human Poldip2 [10].…”

“…Using the SWISS-MODEL program, we determined a model structure of rat Poldip2 (Fig. 1C) based on the recent crystallographic structure of human Poldip2 at 2.8 Å resolution [10]. The resulting structure model of rat Poldip2 was superimposable with the human Poldip2 structures (PDB code 6Z9C and 6ZLX; see Fig.…”

“…The mitochondrial signal is important for the interaction of Polidp2 with PrimPol for DNA synthesis in mitochondria [8] but the truncated form of Poldip2, depleted of its mitochondrial signal has been shown to interact with a plethora of proteins through its two protein-protein interacting domains, the N-terminus YccV-like and the C-terminus DUF525 domains [9], as highlighted by the recent structures of Poldip2 showing the ability of these domains to interact with PrimPol, PCNA [10] and CLPX [11]. The YccV-like domain, highly conserved in eukaryotes, is found in particular in the HspQ (Heat shock protein Q) protein and is also involved in the binding of hemimethylated DNA and the regulation of dnaA gene expression [12][13][14].…”

Self-Sustained Regulation or Self-Perpetuating Dysregulation: ROS-dependent HIF-YAP-Notch Signaling as a Double-Edged Sword on Stem Cell Physiology and Tumorigenesis

“…NOX4 also interacts with a mitochondrial protein, polymerase δ-interacting protein 2 (Poldip2), to increase its activity ( Lyle et al, 2009 ). Poldip2 is a molecule interacting with DNA polymerase δ p50 subunit and with the proteins constituting the mitochondrial DNA nucleoid, through which NOX4 activity is associated with the TCA cycle and metabolic reprogramming ( Andjongo et al, 2021 ; Kulik et al, 2021 ). In fact, metabolism-related cytokines, such as insulin and IGF-1, are known to increase NOX4 expression ( Meng et al, 2008 ; Schroder et al, 2009 ; Kim et al, 2012 ).…”

The polymerase δ-interacting protein family and their emerging roles in diseases

“…POLDIP2 can be expressed in the mitochondria, nucleus, cell membrane, and mitotic spindle. POLDIP2 has two independent domains: the YccV domain at the N-terminal (residues 64–186) and the DUF525 domain at the C-terminal (residues 231–368) ( Figure 2A ) ( 4 , 5 ). An assessment of the POLDIP2 sequence revealed three supposed PCNA binding motifs between residues 81–88, 151–158, and 193–200 ( 6 ).…”