Modular organization of FDH: Exploring the basis of hydrolase catalysis.

An abundant enzyme of liver cytosol, 10-formyltetrahydrofolate dehydrogenase (FDH), is an interesting example of a multidomain protein. It consists of two functionally unrelated domains, an aldehyde dehydrogenase-homologous domain and a folate-binding hydrolase domain, which are connected by an approximately 100-residue linker. The amino-terminal hydrolase domain of FDH (Nt-FDH) is a homolog of formyl transferase enzymes that utilize 10-formyl-THF as a formyl donor.

Disruption of a calmodulin central helix-like region of 10-formyltetrahydrofolate dehydrogenase impairs its dehydrogenase activity by uncoupling the functional domains.

10-Formyltetrahydrofolate dehydrogenase (FDH) is composed of three domains and possesses three catalytic activities but has only two catalytic centers. The amino-terminal domain (residue 1-310) bears 10-formyltetrahydrofolate hydrolase activity, the carboxyl-terminal domain (residue 420-902) bears an aldehyde dehydrogenase activity, and the full-length FDH produces 10-formyltetrahydrofolate dehydrogenase activity. The intermediate linker (residues 311-419) connecting the two catalytic domains does not contribute directly to the enzyme catalytic centers but is crucial for 10-formyltetrahydrofolate dehydrogenase activity.

Amyloid fibril formation of the mouse V(L) domain at acidic pH.

The recombinant V(L) domain that represents the variable part of the light chain (type kappa) of mouse monoclonal antibody F11 directed against human spleen ferritin was found to form amyloid fibrils at acidic pH as evidenced by electron microscopy, thioflavin T binding, and apple-green birefringence after Congo red staining. This is the first demonstration of amyloid fibril formation of the mouse V(L) domain. To understand the mechanism of acidic pH-induced amyloid fibril formation, conformational changes of the V(L) domain were studied by one-dimensional NMR, differential scanning calorimetry, analytical ultracentrifugation, hydrophobic dye binding, far-UV circular dichroism, and tryptophan fluorescence.