Structural elucidation of 5-deoxyribulose-1-phosphate aldolase in a unique 5-deoxyribose metabolism pathway

Qiang Li and Steven D. Bruner

Departments of ‡Chemistry, University of Florida, Gainesville, FL 32611

Radical S-adenosylmethionine (SAM) enzymes utilize a [4Fe−4S] cluster and SAM to initiate a diverse set of complex reactions, in most of the cases via generation of a 5′-deoxyadenosyl radical (dAdo•) intermediate. In the catalytic cycle of a radical SAM, the active intermediate dAdo• abstracts a hydrogen atom and 5'-deoxyadenosine (DOA) is generated as a byproduct. The accumulation of DOA can lead to inhibition of radical SAM enzymes themselves. The metabolism of DOA is known to proceed through 5-deoxyribose via a nucleosidase. However, the fate of 5-deoxyribose is still unclear. Here we propose a pathway for disposal of 5-deoxyribose, which proceeds through a kinase, isomerase and aldolase. Presented, the structure of metal-dependent aldolase, catalyzing the conversion of 5-deoxyribulose-1-phosphate into DHAP and acetaldehyde is described. Aldolase crystals diffract in space group P42₁2, and the structure was determined using the L-fuculose-1-phosphate aldolase from Streptococcus pneumoniae (PDB entry 4C24, 53% sequence identity) as a starting model for molecular replacement. The final model was refined to 1.55 Å resolution with one monomer per asymmetric unit. The structure is composed of a six-stranded antiparallel β-sheet core enfolded by six α-helices of varying lengths. The active site is located on the surface of enzyme with a metal atom coordinated with His95, His97, His157 and Glu76. We calculated different angles in coordination, it shows a trigonal pyramidal shape, which is corresponding to the coordination of manganese (II) ion. EPR results show a relatively strong manganese (II) signal in the ~3,500G range. The crystal structure reveals a novel active site architecture containing a Mn²⁺ ion and defines a unique aldolase structure for dispatching a ubiquitous and unwanted metabolic byproduct.