title: Structural basis for a bispecific NADP+ and CoA binding site in an archaeal malonyl-coenzyme A reductase.
abstract: Autotrophic members of the Sulfolobales (crenarchaeota) use the 3-hydroxypropionate/4-hydroxybutyrate cycle to assimilate CO2 into cell material. The product of the initial acetyl-CoA carboxylation with CO2, malonyl-CoA, is further reduced to malonic semialdehyde by an NADPH-dependent malonyl-CoA reductase (MCR); the enzyme also catalyzes the reduction of succinyl-CoA to succinic semialdehyde onwards in the cycle. Here, we present the crystal structure of Sulfolobus tokodaii malonyl-CoA reductase in the substrate-free state and in complex with NADP(+) and CoA. Structural analysis revealed an unexpected reaction cycle in which NADP(+) and CoA successively occupy identical binding sites. Both coenzymes are pressed into an S-shaped, nearly superimposable structure imposed by a fixed and preformed binding site. The template-governed cofactor shaping implicates the same binding site for the 3'- and 2'-ribose phosphate group of CoA and NADP(+), respectively, but a different one for the common ADP part: the β-phosphate of CoA aligns with the α-phosphate of NADP(+). Evolution from an NADP(+) to a bispecific NADP(+) and CoA binding site involves many amino acid exchanges within a complex process by which constraints of the CoA structure also influence NADP(+) binding. Based on the paralogous aspartate-β-semialdehyde dehydrogenase structurally characterized with a covalent Cys-aspartyl adduct, a malonyl/succinyl group can be reliably modeled into MCR and discussed regarding its binding mode, the malonyl/succinyl specificity, and the catalyzed reaction. The modified polypeptide surrounding around the absent ammonium group in malonate/succinate compared with aspartate provides the structural basis for engineering a methylmalonyl-CoA reductase applied for biotechnical polyester building block synthesis.
authors: Max-Planck-Institut für Biophysik, D-60438 Frankfurt am Main, Germany.
authors: Ulrike
authors: U
authors: Demmer
authors: Eberhard
authors: E
authors: Warkentin
authors: Ankita
authors: A
authors: Srivastava
authors: Daniel
authors: D
authors: Kockelkorn
authors: Markus
authors: M
authors: Pötter
authors: Achim
authors: A
authors: Marx
authors: Georg
authors: G
authors: Fuchs
authors: Ulrich
authors: U
authors: Ermler
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry*; D001665:Binding Sites; D003065:Coenzyme A / Q000737:chemistry*; D009249:NADP / Q000737:chemistry*; D010088:Oxidoreductases / Q000737:chemistry*; D013329:Structure-Activity Relationship; D016944:Sulfolobus / Q000201:enzymology*
chemical_list: D019843:Archaeal Proteins; D009249:NADP; D010088:Oxidoreductases; C454539:malonyl-Coa reductase; D003065:Coenzyme A