A2RJT9 (PYRDA_LACLM) Reviewed, UniProtKB/Swiss-Prot

Names and origin

Protein names

Recommended name:
Dihydroorotate dehydrogenase A (fumarate)
Short name=DHOD A
Short name=DHODase A
Short name=DHOdehase A
EC=1.3.98.1

Gene names

Name:	pyrDA
Ordered Locus Names:	llmg_0952

Organism

Lactococcus lactis subsp. cremoris (strain MG1363) [Complete proteome] [HAMAP]

Taxonomic identifier

416870 [NCBI]

Taxonomic lineage

Bacteria › Firmicutes › Lactobacillales › Streptococcaceae › Lactococcus

Protein attributes

Sequence length	311 AA.
Sequence status	Complete.
Protein existence	Evidence at protein level

General annotation (Comments)

Function	Catalyzes the conversion of dihydroorotate to orotate with fumarate as the electron acceptor. Molecular oxygen can replace fumarate in vitro, but can not use NAD+ as an electron acceptor. Ref.1
Catalytic activity	(S)-dihydroorotate + fumarate = orotate + succinate. Ref.1
Cofactor	Binds 1 FMN per subunit. Ref.4 Ref.5
Pathway	Pyrimidine metabolism; UMP biosynthesis via de novo pathway. HAMAP MF_00224
Subunit structure	Homodimer. Ref.4
Subcellular location	Cytoplasm By similarity HAMAP MF_00224.
Sequence similarities	Belongs to the dihydroorotate dehydrogenase family. Type 1 subfamily.
Biophysicochemical properties	Kinetic parameters: KM=18 µM for dihydroorotate Ref.6 KM=250 µM for fumarate Vmax=18.6 µmol/min/mg enzyme

Ontologies

Keywords
Biological process	Pyrimidine biosynthesis
Cellular component	Cytoplasm
Ligand	FMN Flavoprotein
Molecular function	Oxidoreductase
Technical term	3D-structure Complete proteome
Gene Ontology (GO)
Biological process	'de novo' pyrimidine base biosynthetic process Inferred from electronic annotation. Source: InterPro UMP biosynthetic process Inferred from electronic annotation. Source: InterPro
Cellular component	cytoplasm Inferred from electronic annotation. Source: UniProtKB-SubCell
Molecular function	dihydroorotate oxidase activity Inferred from electronic annotation. Source: InterPro
Complete GO annotation...

Sequence annotation (Features)

Feature key

Position(s)

Length

Description

Graphical view

Feature identifier

Molecule processing

Chain

1 – 311

311

Dihydroorotate dehydrogenase A (fumarate) HAMAP MF_00224

PRO_0000285239

Regions

Nucleotide binding

43 – 44

2

FMN HAMAP MF_00224

Nucleotide binding

249 – 250

2

FMN HAMAP MF_00224

Nucleotide binding

271 – 272

2

FMN HAMAP MF_00224

Region

67 – 71

5

Substrate binding HAMAP MF_00224

Region

193 – 194

2

Substrate binding HAMAP MF_00224

Sites

Active site

130

1

Nucleophile

Binding site

19

1

FMN

Binding site

43

1

Substrate

Binding site

127

1

FMN

Binding site

127

1

Substrate

Binding site

164

1

FMN

Binding site

192

1

FMN; via carbonyl oxygen

Binding site

221

1

FMN; via amide nitrogen

Experimental info

Mutagenesis

43

1

K → A: More than 500-fold reduction of enzymatic activity with oxygen or DCIP as electron acceptor. Ref.3

Mutagenesis

43

1

K → E: 40-fold reduction of enzymatic activity with oxygen as electron acceptor; more than 120-fold reduction with DCIP as electron acceptor. Ref.3

Mutagenesis

50

1

R → E: Steady state kinetics comparable to wild-type; 3-fold decrease in flavin bleaching rate. Ref.6

Mutagenesis

56

1

P → A: More than 500-fold reduction of enzymatic activity. Ref.6

Mutagenesis

57

1

R → A: More active than wild-type, especially with 2,6-dichloroindophenol as electron acceptor. Ref.6

Mutagenesis

67

1

N → A: 100-fold lower affinity for dihydroorotate. Ref.6

Mutagenesis

127

1

N → A: 70-fold lower affinity for dihydroorotate; inhibited by milimolar concentrations of fumarate. Ref.6

Mutagenesis

129

1

S → A: 6-fold lower enzymatic activity with fumarate as electron acceptor. Ref.6

Mutagenesis

130

1

C → A or S: Almost total loss of activity with oxygen or 2,6-dichloroindophenol as electron acceptor. Ref.3

Mutagenesis

131

1

P → A: 4.5-fold lower enzymatic activity with fumarate and 2,6-dichloroindophenol as electron acceptor. Ref.6

Mutagenesis

132

1

N → A: 54-fold reduction of enzymatic activity with oxygen as electron acceptor; more than 250-fold reduction with 2,6-dichloroindophenol as electron acceptor. Ref.3 Ref.6

Mutagenesis

136

1

K → A: Slightly higher affinity to dihydroorotate; 2-fold decrease in flavin bleaching rate. Ref.6

Mutagenesis

164

1

K → A: Almost total loss of activity with oxygen or 2,6-dichloroindophenol as electron acceptor. Ref.3

Mutagenesis

193

1

N → A: 500-fold lower affinity for dihydroorotate; inhibited by milimolar concentrations of fumarate. Ref.6

Mutagenesis

213

1

K → A: 7-fold decrease in enzymatic activity; 50-fold decrease in flavin bleaching rate. Ref.6

Secondary structure

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311

Helix Strand Turn

Details...

Beta strand

4 – 6

3

Beta strand

9 – 17

9

Helix

26 – 34

9

Beta strand

58 – 61

4

Beta strand

64 – 67

4

Helix

76 – 89

14

Beta strand

92 – 94

3

Beta strand

97 – 100

4

Helix

105 – 117

13

Beta strand

122 – 128

7

Beta strand

133 – 135

3

Helix

139 – 141

3

Helix

143 – 153

11

Turn

154 – 156

3

Beta strand

161 – 165

5

Helix

171 – 181

11

Beta strand

188 – 191

4

Beta strand

195 – 199

5

Turn

203 – 206

4

Beta strand

207 – 209

3

Helix

212 – 215

4

Beta strand

216 – 221

6

Helix

222 – 224

3

Helix

225 – 236

12

Beta strand

243 – 250

8

Helix

254 – 263

10

Beta strand

266 – 270

5

Helix

272 – 277

6

Helix

281 – 296

16

Helix

301 – 303

3

Turn

304 – 306

3

Sequences

	Sequence		Length	Mass (Da)
	A2RJT9 [UniParc]. Last modified March 6, 2007. Version 1. Checksum: 30157E3C2791CDD7	FASTA	311	34,210
	10 20 30 40 50 60 MLNTTFANAK FANPFMNASG VHCMTIEDLE ELKASQAGAY ITKSSTLEKR EGNPLPRYVD 70 80 90 100 110 120 LELGSINSMG LPNLGFDYYL DYVLKNQKEN AQEGPIFFSI AGMSAAENIA MLKKIQESDF 130 140 150 160 170 180 SGITELNLSC PNVPGKPQLA YDFEATEKLL KEVFTFFTKP LGVKLPPYFD LVHFDIMAEI 190 200 210 220 230 240 LNQFPLTYVN SVNSIGNGLF IDPEAESVVI KPKDGFGGIG GAYIKPTALA NVRAFYTRLK 250 260 270 280 290 300 PEIQIIGTGG IETGQDAFEH LLCGATMLQI GTALHKEGPA IFDRIIKELE EIMNQKGYQS 310 IADFHGKLKS L

References

	« Hide 'large scale' referencesShow 'large scale' references »
[1]	"Two different dihydroorotate dehydrogenases in Lactococcus lactis." Andersen P.S., Jansen P.J.G., Hammer K. J. Bacteriol. 176:3975-3982(1994) [PubMed: 8021180] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA], FUNCTION, CATALYTIC ACTIVITY, SUBSTRATE SPECIFICITY. Strain: MG1363.
[2]	"The complete genome sequence of the lactic acid bacterial paradigm Lactococcus lactis subsp. cremoris MG1363." Wegmann U., O'Connell-Motherway M., Zomer A., Buist G., Shearman C., Canchaya C., Ventura M., Goesmann A., Gasson M.J., Kuipers O.P., van Sinderen D., Kok J. J. Bacteriol. 189:3256-3270(2007) [PubMed: 17307855] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. Strain: MG1363.
[3]	"Active site of dihydroorotate dehydrogenase A from Lactococcus lactis investigated by chemical modification and mutagenesis." Bjoernberg O., Rowland P., Larsen S., Jensen K.F. Biochemistry 36:16197-16205(1997) [PubMed: 9405053] [Abstract] Cited for: MUTAGENESIS OF LYS-43; CYS-130; ASN-132 AND LYS-164. Strain: MG1363.
[4]	"The crystal structure of the flavin containing enzyme dihydroorotate dehydrogenase A from Lactococcus lactis." Rowland P., Nielsen F.S., Jensen K.F., Larsen S. Structure 5:239-252(1997) [PubMed: 9032071] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.0 ANGSTROMS) IN COMPLEX WITH FMN, COFACTOR, SUBUNIT. Strain: MG1363.
[5]	"The crystal structure of Lactococcus lactis dihydroorotate dehydrogenase A complexed with the enzyme reaction product throws light on its enzymatic function." Rowland P., Bjoernberg O., Nielsen F.S., Jensen K.F., Larsen S. Protein Sci. 7:1269-1279(1998) [PubMed: 9655329] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.0 ANGSTROMS) IN COMPLEX WITH FMN AND OROTATE, COFACTOR, REACTION MECHANISM. Strain: MG1363.
[6]	"Lactococcus lactis dihydroorotate dehydrogenase A mutants reveal important facets of the enzymatic function." Noerager S., Arent S., Bjoernberg O., Ottosen M., Lo Leggio L., Jensen K.F., Larsen S. J. Biol. Chem. 278:28812-28822(2003) [PubMed: 12732650] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (1.7 ANGSTROMS) OF WILD-TYPE AND MUTANTS ALA-56; ARG-56; ALA-67; LYS-136 AND GLU-213 IN COMPLEXES WITH FMN AND SUBSTRATE, MUTAGENESIS OF ARG-50; PRO-56; ARG-57; ASN-67; ASN-127; SER-129; PRO-131; ASN-132; LYS-136; ASN-193 AND LYS-213, BIOPHYSICOCHEMICAL PROPERTIES. Strain: MG1363.
+	Additional computationally mapped references.

Cross-references

Sequence databases

EMBL
GenBank
DDBJ

X74206 Genomic DNA. Translation: CAA52279.1.
AM406671 Genomic DNA. Translation: CAL97544.1.

RefSeq

YP_001032274.1. NC_009004.1.

3D structure databases

PDBe
RCSB PDB
PDBj

Entry	Method	Resolution (Å)	Chain	Positions	PDBsum
1DOR	X-ray	2.00	A/B	1-311	[»]
1JQV	X-ray	2.10	A/B	1-311	[»]
1JQX	X-ray	1.70	A/B	1-311	[»]
1JRB	X-ray	1.90	A/B	1-311	[»]
1JRC	X-ray	1.80	A/B	1-311	[»]
1JUB	X-ray	1.40	A/B	1-311	[»]
1JUE	X-ray	1.80	A/B	1-311	[»]
1NFC	model	-	A	1-311	[»]
1OVD	X-ray	2.25	A/B	1-311	[»]
2BSL	X-ray	2.30	A/B	1-311	[»]
2BX7	X-ray	2.04	A/B	1-311	[»]
2DOR	X-ray	2.00	A/B	1-311	[»]

ProteinModelPortal

A2RJT9.

SMR

A2RJT9. Positions 1-311.

ModBase

Search...

Protein-protein interaction databases

STRING

A2RJT9.

Protocols and materials databases

StructuralBiologyKnowledgebase

Search...

Genome annotation databases

GeneID

4798428.

GenomeReviews

Gene locus llmg_0952 in contig AM406671_GR.

KEGG

llm:llmg_0952.

PATRIC

22283022. VBILacLac4574_0970.

Organism-specific databases

CMR

Search...

Phylogenomic databases

eggNOG

COG0167.

HOGENOM

HBG472415.

OMA

QAAAVFN.

ProtClustDB

PRK02506.

Family and domain databases

HAMAP

MF_00224. DHO_dh_type1.
[Tree]

InterPro

IPR013785. Aldolase_TIM.
IPR005720. Dihydroorotate_DH.
IPR024920. Dihydroorotate_DH_1.
IPR012135. Dihydroorotate_DH_1_2.
IPR001295. Dihydroorotate_DH_CS.
[Graphical view]

Gene3D

G3DSA:3.20.20.70. Aldolase_TIM. 1 hit.

KO

K00226.

Pfam

PF01180. DHO_dh. 1 hit.
[Graphical view]

PIRSF

PIRSF000164. DHO_oxidase. 1 hit.

TIGRFAMs

TIGR01037. PyrD_sub1_fam. 1 hit.

PROSITE

PS00911. DHODEHASE_1. 1 hit.
PS00912. DHODEHASE_2. 1 hit.
[Graphical view]

ProtoNet

Search...

Entry information

Entry name

PYRDA_LACLM

Accession

Primary (citable) accession number: A2RJT9
Secondary accession number(s): P54321

Entry history

Integrated into UniProtKB/Swiss-Prot:	May 1, 2007
Last sequence update:	March 6, 2007
Last modified:	January 25, 2012
This is version 36 of the entry and version 1 of the sequence. [Complete history]

Entry status

Reviewed (UniProtKB/Swiss-Prot)

Annotation program

Prokaryotic Protein Annotation Program