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Title: Crystal structures of the naturally fused CS and cytochrome b 5 reductase ( b 5R) domains of Ncb5or reveal an expanded CS fold, extensive CS– b 5R interactions and productive binding of the NAD(P) + nicotinamide ring

Abstract

Ncb5or (NADH-cytochrome b 5oxidoreductase), a cytosolic ferric reductase implicated in diabetes and neurological diseases, comprises three distinct domains, cytochrome b 5( b 5) and cytochrome b 5reductase ( b 5R) 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 b 5(Cyb5A) and b 5R (Cyb5R3) system, is of interest. A high-resolution structure of the b 5domain (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 b 5R 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 b 5R 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 Cyb5R3more » provide insight into the selectivity for docking of their respective b 5redox partners. The structures also afford a structural explanation for the unusual ability of Ncb5or to utilize both NADH and NADPH, and represent the first examples of native, fully oxidized b 5R family members in which the nicotinamide ring of NAD(P) + resides in the active site. Finally, the structures, together with sequence alignments, show that the b 5R domain is more closely related to single-domain Cyb5R proteins from plants, fungi and some protists than to Cyb5R3 from animals.« less

Authors:
; ; ; ; ; ; ; ORCiD logo
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
NIGMS
OSTI Identifier:
1557317
Resource Type:
Journal Article
Journal Name:
Acta Crystallographica. Section D. Structural Biology
Additional Journal Information:
Journal Volume: 75; Journal Issue: 7; Journal ID: ISSN 2059-7983
Publisher:
IUCr
Country of Publication:
United States
Language:
ENGLISH

Citation Formats

Benson, David R., Lovell, Scott, Mehzabeen, Nurjahan, Galeva, Nadezhda, Cooper, Anne, Gao, Philip, Battaile, Kevin P., and Zhu, Hao. Crystal structures of the naturally fused CS and cytochrome b5 reductase (b5R) domains of Ncb5or reveal an expanded CS fold, extensive CS–b 5R interactions and productive binding of the NAD(P)+ nicotinamide ring. United States: N. p., 2019. Web. doi:10.1107/S205979831900754X.
Benson, David R., Lovell, Scott, Mehzabeen, Nurjahan, Galeva, Nadezhda, Cooper, Anne, Gao, Philip, Battaile, Kevin P., & Zhu, Hao. Crystal structures of the naturally fused CS and cytochrome b5 reductase (b5R) domains of Ncb5or reveal an expanded CS fold, extensive CS–b 5R interactions and productive binding of the NAD(P)+ nicotinamide ring. United States. doi:10.1107/S205979831900754X.
Benson, David R., Lovell, Scott, Mehzabeen, Nurjahan, Galeva, Nadezhda, Cooper, Anne, Gao, Philip, Battaile, Kevin P., and Zhu, Hao. Wed . "Crystal structures of the naturally fused CS and cytochrome b5 reductase (b5R) domains of Ncb5or reveal an expanded CS fold, extensive CS–b 5R interactions and productive binding of the NAD(P)+ nicotinamide ring". United States. doi:10.1107/S205979831900754X.
@article{osti_1557317,
title = {Crystal structures of the naturally fused CS and cytochrome b5 reductase (b5R) domains of Ncb5or reveal an expanded CS fold, extensive CS–b 5R interactions and productive binding of the NAD(P)+ nicotinamide ring},
author = {Benson, David R. and Lovell, Scott and Mehzabeen, Nurjahan and Galeva, Nadezhda and Cooper, Anne and Gao, Philip and Battaile, Kevin P. and Zhu, Hao},
abstractNote = {Ncb5or (NADH-cytochrome b5oxidoreductase), a cytosolic ferric reductase implicated in diabetes and neurological diseases, comprises three distinct domains, cytochrome b5(b5) and cytochrome b5reductase (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 b5domain (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 b5redox partners. The structures also afford a structural explanation for the unusual ability of Ncb5or to utilize both NADH and NADPH, and represent the first examples of native, fully oxidized b5R family members in which the nicotinamide ring of NAD(P)+ resides in the active site. Finally, the structures, together with sequence alignments, show that the b5R domain is more closely related to single-domain Cyb5R proteins from plants, fungi and some protists than to Cyb5R3 from animals.},
doi = {10.1107/S205979831900754X},
journal = {Acta Crystallographica. Section D. Structural Biology},
issn = {2059-7983},
number = 7,
volume = 75,
place = {United States},
year = {2019},
month = {6}
}

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