Derived From Human Sulfur Lipase Superfamily Member 2 And The Lens Proteins Crystallin Mu, The Crystal Structure And Biological Function Study

Human thioesterase superfamily member 2 (hTHEM2) belongs to the hotdog-fold enzyme superfamily but its biological function remains unknown. Tissue specific expression in mouse showed that the encoding gene is highly expressed in the kidney. and moderately expressed in the liver, brain, large intestine and small intestine. Small interference RNA silencing in cell line HCT116 shows that the hthem2 gene is essential for the cell sustained proliferation. Immunostaining and GFP-Tag experiments show that hTHEM2 is co-localized with microtubules.Since hTHEM2 exhibits about 20% sequence identity to Escherichia coli PaaI protein, it was proposed to be a thioesterase with a hotdog-fold. We describe the crystallographic structure of recombinant hTHEM2, determined by the single-wavelength anomalous dispersion method at 2.3 A resolution. This structure demonstrates that hTHEM2 indeed contains a hotdog-fold and forms a back-to-back tetramer as other hotdog proteins. Based on structural and sequence conservation, the thioesterase active site in hTHEM2 is predicted. The structure and substrate specificity are most similar to those of the bacterial phenylacetyl-CoA hydrolase. Asp65, located on the central α-helix of subunit B, was shown by site-directed mutagenesis to be essential to catalysis.Human cytosolic 3,5,3'-triiodo-L-thyronine-binding protein, also called Mu_crystallin or CRYM, plays important physiological roles in transporting 3,5,3'-triiodo-L-thyronine (T3) into nuclei and regulating thyroid hormone related gene expression. We report crystal structure of human CRYM bound with NADPH. The structure contains two domains: a Rossemann fold like NADPH binding domain and a dimerization domain. Different conformations of the loop Arg83-His92 have been observed in two monomers of human CRYM in the same asymmetric unit. The peptide bond of Val89-Pro90 is a trans-configuration in one monomer but a cis-configuration in the other. Finally, a putative T3-binding site in human CRYM is proposed based on comparison with other structural homologues.