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Kaposi's sarcoma-associated herpesvirus (KSHV) is a highly infectious human herpesvirus that causes Kaposi's sarcoma. KSHV encodes functional thymidylate synthase, which is a target for anticancer drugs such as raltitrexed or 5-fluorouracil. Thymidylate synthase catalyzes the conversion of 2'-deoxyuridine-5'-monophosphate (dUMP) to thymidine-5'-monophosphate (dTMP) using 5,10-methylenetetrahydrofolate (mTHF) as a co-substrate. The crystal structures of thymidylate synthase from KSHV (apo), complexes with dUMP (binary), and complexes with both dUMP and raltitrexed (ternary) were determined at 1.7 Å, 2.0 Å, and 2.4 Å, respectively. While the ternary complex structures of human thymidylate synthase and E. coli thymidylate synthase had a closed conformation, the ternary complex structure of KSHV thymidylate synthase was observed in an open conformation, similar to that of rat thymidylate synthase. The complex structures of KSHV thymidylate synthase did not have a covalent bond between the sulfhydryl group of Cys219 and C6 atom of dUMP, unlike the human thymidylate synthase. The catalytic Cys residue demonstrated a dual conformation in the apo structure, and its sulfhydryl group was oriented toward the C6 atom of dUMP with no covalent bond upon ligand binding in the complex structures. These structural data provide the potential use of antifolates such as raltitrexed as a viral induced anticancer drug and structural basis to design drugs for targeting the thymidylate synthase of KSHV., Competing Interests: The authors have declared that no competing interests exist.

The arginine repressor (ArgR) is an arginine-dependent transcription factor that regulates the expression of genes encoding proteins involved in the arginine biosynthesis and catabolic pathways. ArgR is a functional homolog of the arginine-dependent repressor/activator AhrC from Bacillus subtilis, and belongs to the ArgR/AhrC family of transcriptional regulators. In this research, we determined the structure of the ArgR (Bh2777) from Bacillus halodurans at 2.41 Å resolution by X-ray crystallography. The ArgR from B. halodurans appeared to be a trimer in a size exclusion column and in the crystal structure. However, it formed a hexamer in the presence of L-arginine in multi-angle light scattering (MALS) studies, indicating the oligomerization state was dependent on the presence of L-arginine. The trimeric structure showed that the C-terminal domains form the core, which was made by inter-subunit interactions mainly through hydrophobic contacts, while the N-terminal domains containing a winged helix-turn-helix DNA binding motif were arranged around the periphery. The arrangement of trimeric structure in the B. halodurans ArgR was different from those of other ArgR homologs previously reported. We finally showed that the B. halodurans ArgR has an arginine-dependent DNA binding property by an electrophoretic mobility shift assay.

Charcot-Marie-Tooth disease (CMT) is a heterogeneous group of peripheral neuropathies with diverse genetic causes. In this study, we identified p.I43N mutation in PMP2 from a family exhibiting autosomal dominant demyelinating CMT neuropathy by whole exome sequencing and characterized the clinical features. The age at onset was the first to second decades and muscle atrophy started in the distal portion of the leg. Predominant fatty replacement in the anterior and lateral compartment was similar to that in CMT1A caused by PMP22 duplication. Sural nerve biopsy showed onion bulbs and degenerating fibers with various myelin abnormalities. The relevance of PMP2 mutation as a genetic cause of dominant CMT1 was assessed using transgenic mouse models. Transgenic mice expressing wild type or mutant (p.I43N) PMP2 exhibited abnormal motor function. Electrophysiological data revealed that both mice had reduced motor nerve conduction velocities (MNCV). Electron microscopy revealed that demyelinating fibers and internodal lengths were shortened in both transgenic mice. These data imply that overexpression of wild type as well as mutant PMP2 also causes the CMT1 phenotype, which has been documented in the PMP22. This report might expand the genetic and clinical features of CMT and a further mechanism study will enhance our understanding of PMP2-associated peripheral neuropathy.