Abstract
The yeast Candida albicans is the most prevalent opportunistic fungal pathogen in humans. Drug resistance among C. albicans isolates poses a common challenge, and overcoming this resistance represents an unmet need in managing this common pathogen. Here, we investigated CDC8, encoding thymidylate kinase (TMPK), as a potential drug target for the management of C. albicans infections. We found that the region spanning amino acids 106-123, namely the Ca-loop of C. albicans TMPK (CaTMPK), contributes to the hyperactivity of this enzyme compared with the human enzyme (hTMPK) and to the utilization of deoxyuridine monophosphate (dUMP)/deoxy-5-fluorouridine monophosphate (5-FdUMP) as a substrate. Notably, expression of CaTMPK, but not of hTMPK, produced dUTP/5-FdUTP-mediated DNA toxicity in budding yeast (Saccharomyces cerevisiae). CRISPR-mediated deletion of this Ca-loop in C. albicans revealed that the Ca-loop is critical for fungal growth and susceptibility to 5-fluorouridine (5-FUrd). Of note, pathogenic and drug-resistant C. albicans clones were similarly sensitive to 5-FUrd, and we also found that CaTMPK is essential for the growth of C. albicans In conclusion, these findings not only identified a target site for the development of CaTMPK-selective drugs, but also revealed that 5-FUrd may have potential utility as drug for managing C. albicans infections.
Highlights
The yeast Candida albicans is the most prevalent opportunistic fungal pathogen in humans
We found that the region spanning amino acids 106 –123, namely the Ca-loop of C. albicans TMPK (CaTMPK), contributes to the hyperactivity of this enzyme compared with the human enzyme and to the utilization of deoxyuridine monophosphate/deoxy-5-fluorouridine monophosphate (5-FdUMP) as a substrate
Members of the TMPK enzyme family have been categorized into type I and type II enzymes
Summary
We targeted an essential enzyme in C. albicans, thereby selectively suppressing fungal growth and overcoming drug resistance. In C. albicans, TMPK (CaTMPK) is encoded by CDC8, but the essentiality of this gene has not been previously characterized. The main catalytic modules of TMPK include the LID region, P-loop, DR motif, and elements involved in dTMP binding. The sequences of the P-loop, DR motif, and TMP-binding elements are highly conserved among TMPK orthologues, whereas the sequences of the LID region that contribute to the closed conformation for catalysis are divergent. Given the key function of TMPK in dTTP synthesis and the sequence divergence between humans and pathogens, this study investigated the potential application of CaTMPK from C. albicans in antifungal drug development
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