Abstract
Putative tumor suppressor ALDH1L1, the product of natural fusion of three unrelated genes, regulates folate metabolism by catalyzing NADP+-dependent conversion of 10-formyltetrahydrofolate to tetrahydrofolate and CO2. Cryo-EM structures of tetrameric rat ALDH1L1 revealed the architecture and functional domain interactions of this complex enzyme. Highly mobile N-terminal domains, which remove formyl from 10-formyltetrahydrofolate, undergo multiple transient inter-domain interactions. The C-terminal aldehyde dehydrogenase domains, which convert formyl to CO2, form unusually large interfaces with the intermediate domains, homologs of acyl/peptidyl carrier proteins (A/PCPs), which transfer the formyl group between the catalytic domains. The 4′-phosphopantetheine arm of the intermediate domain is fully extended and reaches deep into the catalytic pocket of the C-terminal domain. Remarkably, the tetrameric state of ALDH1L1 is indispensable for catalysis because the intermediate domain transfers formyl between the catalytic domains of different protomers. These findings emphasize the versatility of A/PCPs in complex, highly dynamic enzymatic systems.
Highlights
Putative tumor suppressor ALDH1L1, the product of natural fusion of three unrelated genes, regulates folate metabolism by catalyzing NADP+-dependent conversion of 10formyltetrahydrofolate to tetrahydrofolate and CO2
To reveal the domain organization of ALDH1L1 we performed negative-stain EM (NS-EM) of the full-length ligand-free protein (Rattus norvegicus ALDH1L1 produced in insect cells using a baculovirus expression system) as well as of its individually expressed Ct domain, which forms the rigid tetrameric core of the full-length enzyme
We found that the tetrameric aldehyde dehydrogenase module of ALDH1L1, located at the C terminus, forms the rigid core of the enzyme, whereas the N-terminal hydrolase domains assume a continuum of positions apparently constrained mainly by the length of the inter-domain linkers
Summary
Putative tumor suppressor ALDH1L1, the product of natural fusion of three unrelated genes, regulates folate metabolism by catalyzing NADP+-dependent conversion of 10formyltetrahydrofolate to tetrahydrofolate and CO2. The tetrameric state of ALDH1L1 is indispensable for catalysis because the intermediate domain transfers formyl between the catalytic domains of different protomers These findings emphasize the versatility of A/PCPs in complex, highly dynamic enzymatic systems. The regulatory role of ALDH1L1 is linked to its catalytic reaction, the NADP+-dependent conversion of 10-formyltetrahydrofolate (10-fTHF) to tetrahydrofolate (THF) and CO2, which irreversibly removes onecarbon groups from the folate pool, diminishing the anabolic capacity[1,4] (Fig. 1a). The C-terminal domain (Ct, aa 405–902) belongs to the family of aldehyde dehydrogenases (ALDHs), the group of enzymes catalyzing the conversion of a large variety of aldehydes to corresponding acids using NAD+ or NADP+ as the electron acceptor[25].
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