P83686 (NB5R3_PIG) Reviewed, UniProtKB/Swiss-Prot

Names and origin

Protein names

Recommended name:
NADH-cytochrome b5 reductase 3
Short name=B5R
Short name=Cytochrome b5 reductase
EC=1.6.2.2

Alternative name(s):
Diaphorase-1

Gene names

Name:	CYB5R3
Synonyms:	DIA1

Organism

Sus scrofa (Pig) [Reference proteome]

Taxonomic identifier

9823 [NCBI]

Taxonomic lineage

cellular organisms › Eukaryota › Opisthokonta › Metazoa › Eumetazoa › Bilateria › Deuterostomia › Chordata › Craniata › Vertebrata › Gnathostomata › Teleostomi › Euteleostomi › Sarcopterygii › Tetrapoda › Amniota › Mammalia › Theria › Eutheria › Laurasiatheria › Cetartiodactyla › Suina › Suidae › Sus

Protein attributes

Sequence length	272 AA.
Sequence status	Fragment.
Protein existence	Evidence at protein level

General annotation (Comments)

Function	Desaturation and elongation of fatty acids, cholesterol biosynthesis, drug metabolism, and, in erythrocyte, methemoglobin reduction.
Catalytic activity	NADH + 2 ferricytochrome b5 = NAD+ + H+ + 2 ferrocytochrome b5.
Cofactor	FAD. Ref.4
Subunit structure	Component of a complex composed of cytochrome b5, NADH-cytochrome b5 reductase (CYB5R3) and MOSC2. Ref.9
Subcellular location	Endoplasmic reticulum membrane; Lipid-anchor; Cytoplasmic side By similarity. Mitochondrion outer membrane; Lipid-anchor; Cytoplasmic side By similarity. Cytoplasm By similarity. Note: The enzyme exists in two forms, a membrane-bound form on the cytoplasmic side of the endoplasmic reticulum and also on the mitochondrial outer membrane and in soluble form in erythrocytes By similarity. NADH-cytochrome b5 reductase 3 membrane-bound form: Endoplasmic reticulum membrane; Lipid-anchor; Cytoplasmic side By similarity. Mitochondrion outer membrane; Lipid-anchor; Cytoplasmic side By similarity. NADH-cytochrome b5 reductase 3 soluble form: Cytoplasm By similarity.
Sequence similarities	Belongs to the flavoprotein pyridine nucleotide cytochrome reductase family. Contains 1 FAD-binding FR-type domain.

Ontologies

Keywords
Biological process	Cholesterol biosynthesis Cholesterol metabolism Lipid biosynthesis Lipid metabolism Steroid biosynthesis Steroid metabolism Sterol biosynthesis Sterol metabolism
Cellular component	Cytoplasm Endoplasmic reticulum Membrane Mitochondrion Mitochondrion outer membrane
Ligand	FAD Flavoprotein NAD
Molecular function	Oxidoreductase
PTM	Acetylation Lipoprotein Phosphoprotein
Technical term	3D-structure Complete proteome Direct protein sequencing Reference proteome
Gene Ontology (GO)
Biological_process	cholesterol biosynthetic process Non-traceable author statement. Source: UniProtKB drug metabolic process Non-traceable author statement. Source: UniProtKB fatty acid elongation Non-traceable author statement. Source: UniProtKB unsaturated fatty acid biosynthetic process Non-traceable author statement. Source: UniProtKB
Cellular_component	endoplasmic reticulum membrane Inferred from electronic annotation. Source: UniProtKB-SubCell mitochondrial outer membrane Inferred from electronic annotation. Source: UniProtKB-SubCell
Molecular_function	cytochrome-b5 reductase activity, acting on NAD(P)H Inferred from direct assay Ref.3. Source: UniProtKB flavin adenine dinucleotide binding Inferred from direct assay Ref.4. Source: UniProtKB
Complete GO annotation...

Sequence annotation (Features)

Feature key

Position(s)

Length

Description

Graphical view

Feature identifier

Molecule processing

Chain

‹1 – 272

›272

NADH-cytochrome b5 reductase 3

PRO_0000167625

Regions

Domain

11 – 123

113

FAD-binding FR-type

Nucleotide binding

103 – 118

16

FAD By similarity

Nucleotide binding

142 – 177

36

FAD By similarity UniProtKB P07514

Amino acid modifications

Modified residue

13

1

N6-acetyllysine By similarity

Modified residue

14

1

Phosphotyrosine By similarity

Modified residue

91

1

N6-acetyllysine By similarity

Modified residue

101

1

Phosphotyrosine By similarity

Experimental info

Mutagenesis

49

1

H → A: Similar properties to wild-type. Ref.1

Mutagenesis

49

1

H → E: Decreased Kcat values for PB5, ferricyanide and NADH. Increased Km values for all three substrates. Ref.1

Mutagenesis

49

1

H → K: Decreased Kcat and Km values for PB5 and ferricyanide. Increased values for NADH. Ref.1

Mutagenesis

49

1

H → Y: Similar properties to wild-type. Ref.1

Mutagenesis

63

1

R → A or Q: Increased Km values for NADH and increased dissociation constant for NAD(+). Ref.7

Mutagenesis

63

1

R → K: Little effect on Km and KD values. Ref.7

Mutagenesis

65

1

Y → A or F: Protein destabilized, release of FAD accelerated, and Kcat values decreased. Ref.7

Mutagenesis

66

1

T → A: Similar properties to wild-type. Ref.8

Mutagenesis

66

1

T → S: Similar properties to wild-type. Ref.8

Mutagenesis

66

1

T → V: 10% of wild-type Kcat values. Ref.8

Mutagenesis

97

1

K → A: Little effect on absorption or CD spectra. Ref.7

Mutagenesis

97

1

K → R: Little effect on absorption or CD spectra. Ref.7

Mutagenesis

99

1

S → A or V: Increased Km values for NADH and increased dissociation constant for NAD(+). Ref.7

Mutagenesis

99

1

S → T: Little effect on Km and KD values. Ref.7

Mutagenesis

272

1

F → FG: 10% of wild-type Kcat values. Ref.1

Mutagenesis

272

1

Missing: 54% of wild-type Kcat values. Ref.1

Sequence conflict

39

1

E → Q Ref.1

Non-terminal residue

1

1

Secondary structure

1

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272

Helix Strand Turn

Details...

Beta strand

12 – 23

12

Beta strand

26 – 33

8

Beta strand

49 – 56

8

Beta strand

59 – 65

7

Beta strand

79 – 83

5

Beta strand

89 – 91

3

Helix

97 – 104

8

Beta strand

110 – 117

8

Beta strand

119 – 124

6

Beta strand

127 – 130

4

Beta strand

132 – 135

4

Beta strand

137 – 139

3

Beta strand

145 – 151

7

Helix

152 – 154

3

Helix

155 – 167

13

Beta strand

174 – 183

10

Helix

184 – 186

3

Helix

190 – 194

5

Helix

197 – 200

4

Turn

201 – 203

3

Beta strand

204 – 212

9

Beta strand

219 – 222

4

Helix

227 – 232

6

Helix

236 – 238

3

Beta strand

241 – 246

6

Helix

249 – 251

3

Helix

257 – 261

5

Helix

266 – 268

3

Sequences

	Sequence		Length	Mass (Da)
	P83686 [UniParc]. Last modified December 15, 2003. Version 1. Checksum: 16B08682FC365090	FASTA	272	30,831
	10 20 30 40 50 60 STPAITLENP DIKYPLRLID KEVVNHDTRR FRFALPSPEH ILGLPVGQHI YLSARIDGNL 70 80 90 100 110 120 VIRPYTPVSS DDDKGFVDLV IKVYFKDTHP KFPAGGKMSQ YLESMKIGDT IEFRGPNGLL 130 140 150 160 170 180 VYQGKGKFAI RPDKKSSPVI KTVKSVGMIA GGTGITPMLQ VIRAIMKDPD DHTVCHLLFA 190 200 210 220 230 240 NQTEKDILLR PELEELRNEH SARFKLWYTV DRAPEAWDYS QGFVNEEMIR DHLPPPEEEP 250 260 270 LVLMCGPPPM IQYACLPNLE RVGHPKERCF AF

References

[1]	"Systematic mutations of highly conserved His49 and carboxyl-terminal of recombinant porcine liver NADH-cytochrome b5 reductase solubilized domain." Kimura S., Emi Y., Ikushiro S., Iyanagi T. Biochim. Biophys. Acta 1430:290-301(1999) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE, PROTEIN SEQUENCE OF 1-5, MUTAGENESIS OF HIS-49 AND PHE-272, CIRCULAR DICHROISM ANALYSIS. Tissue: Liver.
[2]	"Terminal sequences of lysosome solubilized pig liver cytochrome b5 reductase." Crabb J.W., Tarr G.E., Yasunobu K.T., Iyanagi T., Coon M.J. Biochem. Biophys. Res. Commun. 95:1650-1655(1980) [PubMed] [Europe PMC] [Abstract] Cited for: PROTEIN SEQUENCE OF 1-20. Tissue: Liver.
[3]	"Gastric microsomal NADH-cytochrome b5 reductase: characterization and solubilization." Ghesquier D., Robert J.C., Soumarmon A., Abastado M., Grelac F., Lewin M.J.M. Comp. Biochem. Physiol. 80B:165-169(1985) [PubMed] [Europe PMC] [Abstract] Cited for: CHARACTERIZATION.
[4]	"Specific arrangement of three amino acid residues for flavin-binding barrel structures in NADH-cytochrome b5 reductase and the other flavin-dependent reductases." Nishida H., Inaka K., Miki K. FEBS Lett. 361:97-100(1995) [PubMed] [Europe PMC] [Abstract] Cited for: FAD-BINDING.
[5]	"Redox properties of microsomal reduced nicotinamide adenine dinucleotide-cytochrome b5 reductase and cytochrome b5." Iyanagi T. Biochemistry 16:2725-2730(1977) [PubMed] [Europe PMC] [Abstract] Cited for: POTENTIOMETRIC TITRATION, EPR SPECTROSCOPY.
[6]	"One-electron reduction of hepatic NADH-cytochrome b5 reductase as studied by pulse radiolysis." Kobayashi K., Iyanagi T., Ohara H., Hayashi K. J. Biol. Chem. 263:7493-7499(1988) [PubMed] [Europe PMC] [Abstract] Cited for: ENZYME KINETICS.
[7]	"Effects of flavin-binding motif amino acid mutations in the NADH-cytochrome b5 reductase catalytic domain on protein stability and catalysis." Kimura S., Nishida H., Iyanagi T. J. Biochem. 130:481-490(2001) [PubMed] [Europe PMC] [Abstract] Cited for: MUTAGENESIS OF ARG-63; TYR-65; LYS-97 AND SER-99, CIRCULAR DICHROISM ANALYSIS.
[8]	"Role of Thr(66) in porcine NADH-cytochrome b5 reductase in catalysis and control of the rate-limiting step in electron transfer." Kimura S., Kawamura M., Iyanagi T. J. Biol. Chem. 278:3580-3589(2003) [PubMed] [Europe PMC] [Abstract] Cited for: MUTAGENESIS OF THR-66, ENZYME KINETICS, CIRCULAR DICHROISM ANALYSIS.
[9]	"Identification of the missing component in the mitochondrial benzamidoxime prodrug converting system as a novel molybdenum enzyme." Havemeyer A., Bittner F., Wollers S., Mendel R., Kunze T., Clement B. J. Biol. Chem. 281:34796-34802(2006) [PubMed] [Europe PMC] [Abstract] Cited for: IDENTIFICATION IN A COMPLEX WITH CYTOCHROME B5 AND MOSC2.
[10]	"Crystal structure of NADH-cytochrome b5 reductase from pig liver at 2.4 A resolution." Nishida H., Inaka K., Yamanaka M., Kaida S., Kobayashi K., Miki K. Biochemistry 34:2763-2767(1995) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.4 ANGSTROMS) OF 2-273.
[11]	"Electrostatic properties deduced from refined structures of NADH-cytochrome b5 reductase and the other flavin-dependent reductases: pyridine nucleotide-binding and interaction with an electron-transfer partner." Nishida H., Miki K. Proteins 26:32-41(1996) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.1 ANGSTROMS), 3D-STRUCTURE MODELING OF COMPLEX WITH CYTOCHROME B5.

Cross-references

3D structure databases

PDBe
RCSB PDB
PDBj

Entry	Method	Resolution (Å)	Chain	Positions	PDBsum
1NDH	X-ray	2.10	A	1-272	[»]

ProteinModelPortal

P83686.

SMR

P83686. Positions 3-272.

ModBase

Search...

Protein-protein interaction databases

STRING

9823.ENSSSCP00000000040.

Proteomic databases

PaxDb

P83686.

Protocols and materials databases

StructuralBiologyKnowledgebase

Search...

Phylogenomic databases

eggNOG

COG0543.

HOGENOM

HOG000175005.

HOVERGEN

HBG052580.

OrthoDB

EOG418BNW.

Enzyme and pathway databases

SABIO-RK

P83686.

Family and domain databases

InterPro

IPR017927. Fd_Rdtase_FAD-bd.
IPR001709. Flavoprot_Pyr_Nucl_cyt_Rdtase.
IPR001834. NADH-Cyt_B5_reductase.
IPR008333. OxRdtase_FAD-bd_dom.
IPR001433. OxRdtase_FAD/NAD-bd.
IPR017938. Riboflavin_synthase-like_b-brl.
[Graphical view]

Pfam

PF00970. FAD_binding_6. 1 hit.
PF00175. NAD_binding_1. 1 hit.
[Graphical view]

PRINTS

PR00406. CYTB5RDTASE.
PR00371. FPNCR.

SUPFAM

SSF63380. Riboflavin_synthase_like_b-brl. 1 hit.

PROSITE

PS51384. FAD_FR. 1 hit.
[Graphical view]

ProtoNet

Search...

Other

EvolutionaryTrace

P83686.

Entry information

Entry name

NB5R3_PIG

Accession

Primary (citable) accession number: P83686

Entry history

Integrated into UniProtKB/Swiss-Prot:	December 15, 2003
Last sequence update:	December 15, 2003
Last modified:	April 3, 2013
This is version 80 of the entry and version 1 of the sequence. [Complete history]

Entry status

Reviewed (UniProtKB/Swiss-Prot)

Annotation program

Chordata Protein Annotation Program