Structural basis for a bispecific NADP+ and CoA binding site in an archaeal malonyl-coenzyme A reductase

J Biol Chem. 2013 Mar 1;288(9):6363-70. doi: 10.1074/jbc.M112.421263. Epub 2013 Jan 16.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Archaeal Proteins / chemistry*
  • Binding Sites
  • Coenzyme A / chemistry*
  • NADP / chemistry*
  • Oxidoreductases / chemistry*
  • Structure-Activity Relationship
  • Sulfolobus / enzymology*

Substances

  • Archaeal Proteins
  • NADP
  • Oxidoreductases
  • malonyl-Coa reductase
  • Coenzyme A

Associated data

  • PDB/4DPK
  • PDB/4DPL
  • PDB/4DPM