Structural analyses of PCNA from the fungal pathogen Candida albicans identify three regions with species‐specific conformations

Abstract

An assembly of multiprotein complexes achieves chromosomal DNA replication at the replication fork. In eukaryotes, proliferating cell nuclear antigen (PCNA) plays a vital role in the assembly of multiprotein complexes at the replication fork and is essential for cell viability. PCNA from several organisms, including Saccharomyces cerevisiae, has been structurally characterised. However, the structural analyses of PCNA from fungal pathogens are limited. Recently, we have reported that PCNA from the opportunistic fungal pathogen Candida albicans complements the essential functions of ScPCNA in S. cerevisiae. Still, it only partially rescues the loss of ScPCNA when the yeast cells are under genotoxic stress. To understand this further, herein, we have determined the crystal structure of CaPCNA and compared that with the existing structures of other fungal and human PCNA. Our comparative structural and in‐solution small‐angle X‐ray scattering (SAXS) analyses reveal that CaPCNA forms a stable homotrimer, both in crystal and in solution. It displays noticeable structural alterations in the oligomerisation interface, P‐loop and hydrophobic pocket regions, suggesting its differential function in a heterologous system and avenues for developing specific therapeutics. Databases The PDB and SASBDB accession codes for CaPCNA are https://doi.org/10.2210/pdb7BUP/pdb and SASDHQ9, respectively.

Interdomain connecting loop and J loop structures determine cross-species compatibility of PCNA

“…Although CaPCNA possesses significant IDCL sequence identity and belongs to group-I, contrary to the earlier report, CaPCNA is functionally noncompatible in S. cerevisiae (6,9,11). Our comparative structural analyses revealed that CaPCNA displayed noticeable structural alterations in the hydrophobic pocket, J loop, and P loop regions (9).…”

“…Despite our repeated efforts with varying concentrations of ScPCNA and T2AA, no significant heat exchange was observed, suggesting a weak or undetectable interaction between ScPCNA and T2AA. In our earlier study, we had observed that while human PCNA interacts quite strongly with T2AA, CaPCNA failed to show such binding, suggesting that T2AA has a higher affinity for group-II PCNA than for group-I PCNA (9). Although T2AA binds to NcPCNA, it does not inhibit NcPCNA functions in yeast cells, probably due to its lower affinity (M) than the human PCNA (nM) (Fig.…”

“…Irrespective of sequence homology, the conformations of these loop regions are distinct among different species and could be responsible for species-specificity. The IDCL loop, P loop, and J loop exhibit considerable flexibility and adopt different states (9). Usually, due to high flexibility, IDCLs are unstructured in some of the protomers of the various available PCNA trimer structures.…”

“…PCNA has been characterized in human, rat, budding, and fission yeasts, protozoan, flies, Arabidopsis, and many archaeal species (2,6). The crystal structures of PCNA from some of these organisms have also been determined (7)(8)(9)(10). A typical eukaryotic PCNA consists of three protomers to form a toroidal-shaped structure with pseudo-six fold symmetry that encircles the DNA double helix.…”

“…To understand species specificity of PCNA and its functional compatibility across fungal species in greater detail, we have characterized PCNA from N. crassa and compared it with PCNA from S. cerevisiae, and the pathogenic yeasts C. albicans and A. fumigatus (6,9,10), two representatives from each compatible groups. Although CaPCNA possesses significant IDCL sequence identity and belongs to group-I, contrary to the earlier report, CaPCNA is functionally noncompatible in S. cerevisiae (6,9,11). Our comparative structural analyses revealed that CaPCNA displayed noticeable structural alterations in the hydrophobic pocket, J loop, and P loop regions (9).…”