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Protein Eng Des Sel. 2017 Mar 1;30(3):253-261. doi: 10.1093/protein/gzx004.

Understanding the molecular mechanism of substrate channeling and domain communication in protozoal bifunctional TS-DHFR.

Author information

1
Departments of Pharmacology and Molecular Biophysics and Biochemistry, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520-8066,USA.

Abstract

Most species, such as humans, have monofunctional forms of thymidylate synthase (TS) and dihydrofolate reductase (DHFR) that are key folate metabolism enzymes making critical folate components required for DNA synthesis. In contrast, several parasitic protozoa, including Leishmania major (Lm), Plasmodium falciparum (Pf), Toxoplasma gondii (Tg) and Cryptosporidium hominis (Ch), contain a unique bifunctional thymidylate synthase-dihydrofolate reductase (TS-DHFR) having the two sequential catalytic activities contained on a single polypeptide chain. It has been suggested that the bifunctional nature of the two catalytic activities may enable substrate channeling. The 3D structures for each of these enzymes reveals distinct features for each species. While three of the four species (Pf, Tg and Ch) contain a junctional region linking the two domains, this is lacking in Lm. The Lm and Pf contain N-terminal amino acid extensions. A multidisciplinary approach using structural studies and transient kinetic analyses combined with mutational analysis has investigated the roles of these unique structural features for each enzyme. Additionally, the possibility of substrate channeling behavior was explored. These studies have identified unique, functional regions in both the TS and DHFR domains that govern efficient catalysis for each species. Surprisingly, even though there are structural similarities among the species, each is regulated in a distinct manner. This structural and mechanistic information was also used to exploit species-specific inhibitor design.

KEYWORDS:

bifunctional; dihydrofolate; reductase; synthase; thymidylate

PMID:
28338744
PMCID:
PMC6438133
DOI:
10.1093/protein/gzx004
[Indexed for MEDLINE]
Free PMC Article

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