Cryptococcus neoformans as a Model Fungal Pathogen [chapter]

Klaus Lengeler, Joseph Heitman
2002 Molecular Biology of Fungal Development  
CK2α is a constitutively active and highly conserved serine/threonine protein kinase that is involved in the regulation of key cellular metabolic pathways and associated with a variety of tumours and cancers. The most well-known CK2α inhibitor is the human clinical trial candidate CX-4945, which has recently shown to exhibit not only anti-cancer, but also anti-fungal properties. This prompted us to work on the CK2α orthologue, Cka1, from the pathogenic fungus Cryptococcus neoformans, which
more » ... formans, which causes life-threatening systemic cryptococcosis and meningoencephalitis mainly in immunocompromised individuals. At present, treatment of cryptococcosis remains a challenge due to limited anticryptococcal therapeutic strategies. Hence, expanding therapeutic options for the treatment of the disease is highly clinically relevant. Herein, we report the structures of Cka1-AMPPNP-Mg 2+ (2.40 Å) and Cka1-CX-4945 (2.09 Å). Structural comparisons of Cka1-AMPPNP-Mg 2+ with other orthologues revealed the dynamic architecture of the N-lobe across species. This may explain for the difference in binding affinities and deviations in protein-inhibitor interactions between Cka1-CX-4945 and human CK2α-CX-4945. Supporting it, in vitro kinase assay demonstrated that CX-4945 inhibited human CK2α much more efficiently than Cka1. Our results provide structural insights into the design of more selective inhibitors against Cka1. Protein kinase CK2 (formerly known as casein kinase 2) is a constitutively active, ubiquitous and highly conserved serine/threonine kinase complex that lies within the CMGC group of protein kinases 1,2 . Given the importance of protein phosphorylation for a number of cellular processes, protein kinases are often regarded as attractive therapeutic targets and central interests of structure-based drug design 3 . Notably, CK2 is a highly pleiotropic enzyme with more than 300 substrates reported 1 . Due to its vast array of cellular targets, CK2 is involved in a diversity of complex cellular mechanisms relating to the maintenance of cell growth and cell viability 4 . More specifically, CK2 regulates several oncogenic intracellular signal transduction pathways, including the PI3K/AKT, NF-κB, and JAK-STAT pathways, which in turn alter gene expression extensively and promote cell proliferation, survival, and angiogenesis 5 . CK2 is a heterotetrameric enzyme comprised of two catalytic subunits (CK2α) and a central dimer of regulatory subunits (CK2β) 6 . Studies have shown that the subunits consist of multiple isoforms in different organisms. In humans, three isoforms of the catalytic subunit have been discovered, namely CK2α, CK2α′ and CK2α′′, Structure of the cnCka1-CX-4945 complex. To gain an insight into the binding affinity between cnCka1 and CX-4945 for the development of selective anti-fungal drugs, the near full-length structure of cnCka1 in complex with CX-4945 (excluding residues 1-4 and residues 334-338 which are not visible in the electron density map) was subsequently solved and refined to 2.09 Å (Fig. 2a) . Similar to the AMPPNP-Mg 2+ bound structure, CX-4945 binds at the hydrophobic catalytic cleft (Fig. 2b) and interacts with the residues mostly through hydrophobic interactions (Fig. 2d) . Additionally, hydrogen bonds are formed between Lys67 and CX-4945, as well as the backbones of Val115 and Asp174 with CX-4945, and an electrostatic interaction is formed between the backbone of Arg46 with the chloride atom of CX-4945. In general, due to the ring structures of the CX-4945 molecule, the inhibitor bound complex (Fig. 2d ) contains more hydrophobic interactions as compared to the AMPPNP-Mg 2+ bound structure (Fig. 1d) .
doi:10.1201/9780203910719.ch19 fatcat:a3tbkqe3sbfyneg73cde57ujza