Study of the Individual Cytochrome b5 and Cytochrome b5 Reductase Domains of Ncb5or Reveals a Unique Heme Pocket and a Possible Role of the CS Domain

Abstract

(Fig. 1A). Several natural fusion proteins are known that contain a cytochrome b 5 domain and a second redox-active domain. Examples include sulfite oxidase (2-4) and ⌬5 and ⌬6 fatty acid desaturases (5, 6) in animals, nitrate reductase in algae (7) and plants (8, 9), and flavocytochrome b 2 or lactate dehydrogenase (10) and ⌬9 fatty acid desaturase (11) in bakers' yeast. However, Ncb5or is the only member of the cytochrome b 5 superfamily known to contain three distinct domains. The function of the non-redox-active CS domain is presently unknown, although its primary sequence is distantly homologous to those in human heat shock protein 20 (HSP20, a co-chaperone of HSP90) and other CS family members (12).The cytochrome b 5 family includes two isoforms in vertebrates, one anchored to the membrane of the endoplasmic reticulum (Cyb5A) and the other anchored to the outer mitochondrial membrane (Cyb5B) (13). Membrane anchoring in Cyb5A and Cyb5B is accomplished by a hydrophobic C-terminal domain. The polar heme-binding domains of Cyb5A and Cyb5B have virtually identical folds with secondary structure elements occurring in the order ␤1-␣1-␤4-␤3-␣2-␣3-␤5-␣4-* This work was supported, in whole or in part, by National Institutes of Health Grant RO1 DK067355 (to H. Franklin Bunn and H. Z.) and by National Institutes of Health Centers of Biomedical Research Excellence-Protein Structure and Function award 5P20 RR17708 (to the University of Kansas, R. P. Hanzlik, P. I.). This work was also supported by American Heart Association Grant-in-Aid 0755879T (to B. R. G.

Lipotoxicity in the liver

“…Hepatocytes of Ncb5or knockout mice have an increased sensitivity to SFA-induced damage, which can be at least partly attributed to a lower SCD-specific activity in these cells [27] . This observation strongly supports the hypothesized functional link between SCD1 and Ncb5or [8] . Some natural human mutations of NCB5OR have been reported to remarkably decrease the level of active enzyme due to enhanced proteasomal degradation [28] ; therefore, the potential role of this gene in human pathology deserves further investigation.…”

“…to participate in fatty acid desaturation [8] . Membrane lipid saturation is regulated through the activity of various lysophospolipid acyltransferase enzymes exhibiting different acyl-CoA specificities.…”

Ncb5or Deficiency Increases Fatty Acid Catabolism and Oxidative Stress

“…Ncb5or readily reduces a variety of artificial substrates in vitro in the presence of excess NADH or NADPH, as does the Cyb5A⅐Cyb5R3 complex (1,11,12). Ncb5or is a more potent electron donor than Cyb5A because of the lower redox potential (E 0 ϭ Ϫ108 mV) in the Ncb5or heme center (11) and its unique heme environment (13). Of four recognized missense mutations in the human Ncb5or gene, two (Q187R and H223R) are found in a cohort with non-autoimmune diabetes (14).…”

Role of Protein-Protein Interactions in Metabolism: Genetics, Structure, Function

“…In this complex, it became clear that shortening the CPR hinge region leads to a protein that favors the open conformation and promotes association with HO (Sugishima et al, 2014). These studies demonstrate the utility of combining the power of Sevrioukova et al, 1997Cupp-Vickery et al, 2000;Hall et al, 2001;Podust et al, 2001;Yoshinari et al, 2001;Williams et al, 2003;Grahn et al, 2006;He et al, 2006;Deng et al, 2010;Wilderman et al, 2012;Hiruma et al, 2013;Tripathi et al, 2013;Peng et al, 2014;Sugishima et al, 2014;Basudhar et al, 2015;Reed and Backes, 2017;Gill, 1983;Kakuta et al, 1997;Meech and Mackenzie, 1997;Argiriadi et al, 1999;Binda et al, 2001Binda et al, , 2011…”

“…In an X-ray crystallography study comparing the b 5 -like protein, Ncb5or, with b 5 , Deng et al determined that the positioning of the second histidine heme ligand (His 112) in b 5 is critical for efficient electron transfer, both from b 5 reductase and to the (CYP) electron transfer partner (Figure 1; Deng et al, 2010). These are just a few of the many examples available where X-ray crystallography has been used to interrogate CYP and electron transfer partner interactions.…”