Crystal structures of the naturally fused CS and cytochrome b5 reductase (b5R) domains of Ncb5or reveal an expanded CS fold, extensive CS – b5R interactions and productive binding of the NAD(P)+ nicotinamide ring

Ncb5or (NADH-cytochrome b5 oxidoreductase), a cytosolic ferric reductase implicated in diabetes and neurological diseases, comprises three distinct domains, cytochrome b5 (b5) and cytochrome b5 reductase (b5R) domains separated by a CHORD – Sgt1 (CS) domain, and a novel 50-residue N-terminal region. Understanding how interdomain interactions in Ncb5or facilitate the shuttling of electrons from NAD(P)H to heme, and how the process compares with the microsomal b5 (Cyb5A) and b5R (Cyb5R3) system, is of interest. A high-resolution structure of the b5 domain (PDB entry 3lf5) has previously been reported, which exhibits substantial differences in comparison to Cyb5A. The structural characterization of a construct comprising the naturally fused CS and b5R domains with bound FAD and NAD+ (PDB entry 6mv1) or NADP+ (PDB entry 6mv2) is now reported. The structures reveal that the linker between the CS and b5R cores is more ordered than predicted, with much of it extending the β -sandwich motif of the CS domain. This limits the flexibility between the two domains, which recognize one another via a short β -sheet motif and a network of conserved side-chain hydrogen bonds, salt bridges and cation – π interactions. Notable differences in FAD – protein interactions in Ncb5or and Cyb5R3 provide insight into the selectivity for docking of their respective b5 redox partners. The structures also afford a structural explanation for the unusual ability of Ncb5or to utilize both NADH and N...
Source: Acta Crystallographica Section D - Category: Biochemistry Authors: Tags: crystal structure CHORD – Sgt1 domain CS domain FAD NAD NADP Ncb5or research papers Source Type: research