P06632 (DKGA_CORSC) Reviewed, UniProtKB/Swiss-Prot

Names and origin

Protein names

Recommended name:
2,5-diketo-D-gluconic acid reductase A
Short name=2,5-DKG reductase A
Short name=2,5-DKGR A
Short name=25DKGR-A
EC=1.1.1.346

Alternative name(s):
AKR5C

Gene names

Name:

dkgA

Organism

Corynebacterium sp. (strain ATCC 31090)

Taxonomic identifier

268952 [NCBI]

Taxonomic lineage

cellular organisms › Bacteria › Actinobacteria › Actinobacteria › Actinobacteridae › Actinomycetales › Corynebacterineae › Corynebacteriaceae › Corynebacterium

Protein attributes

Sequence length	278 AA.
Sequence status	Complete.
Sequence processing	The displayed sequence is further processed into a mature form.
Protein existence	Evidence at protein level

General annotation (Comments)

Function	Catalyzes the reduction of 2,5-diketo-D-gluconic acid (25DKG) to 2-keto-L-gulonic acid (2KLG). 5-keto-D-fructose and dihydroxyacetone can also serve as substrates. 25DKGR-A exhibits a greater selectivity for the substrate and higher thermal stability than 25DKGR-B.
Catalytic activity	2-dehydro-L-gulonate + NADP+ = 2,5-didehydro-D-gluconate + NADPH.
Enzyme regulation	Inhibited by Zn2+, Fe3+, Cu2+ and Ni2+.
Subunit structure	Monomer.
Subcellular location	Cytoplasm.
Biotechnological use	Introduced by genetic manipulation and expressed in Erwinia herbicola by Anderson et al. The resultant organism is able to convert in a single fermentative step D-glucose into 2-keto-L-gulonic acid, a key precursor in the industrial production of L-ascorbic acid (vitamin C). However, the technology still needs some working on and is not used in commercial production at present.
Miscellaneous	Modeling study indicates that the active site may not be optimized for 2,5-diketo-D-gluconic acid.
Sequence similarities	Belongs to the aldo/keto reductase family.
Biophysicochemical properties	Kinetic parameters: KM=26 mM for 2,5-diketo-D-gluconate Ref.2 KM=10 µM for NADPH pH dependence: Optimum pH is 6.4. Active over a broad pH range.

Ontologies

Keywords
Biological process	Ascorbate biosynthesis
Cellular component	Cytoplasm
Ligand	NADP
Molecular function	Oxidoreductase
Technical term	3D-structure Direct protein sequencing
Gene Ontology (GO)
Biological_process	L-ascorbic acid biosynthetic process Inferred from electronic annotation. Source: UniProtKB-KW
Cellular_component	cytoplasm Inferred from electronic annotation. Source: UniProtKB-SubCell
Molecular_function	oxidoreductase activity Inferred from electronic annotation. Source: UniProtKB-KW
Complete GO annotation...

Sequence annotation (Features)

Feature key

Position(s)

Length

Description

Graphical view

Feature identifier

Molecule processing

Initiator methionine

1

1

Removed Ref.1 Ref.2

Chain

2 – 278

277

2,5-diketo-D-gluconic acid reductase A

PRO_0000124597

Regions

Nucleotide binding

188 – 242

55

NADP

Sites

Active site

50

1

Proton donor

Binding site

108

1

Substrate

Experimental info

Mutagenesis

22

1

F → Y: Tighter binding of NADH.

Mutagenesis

232

1

K → G: Tighter binding of NADH.

Mutagenesis

238

1

R → H: Tighter binding of NADH.

Mutagenesis

272

1

A → G: Tighter binding of NADH.

Secondary structure

1

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278

Helix Strand Turn

Details...

Beta strand

5 – 7

3

Beta strand

13 – 17

5

Helix

26 – 28

3

Helix

29 – 39

11

Beta strand

43 – 45

3

Helix

47 – 49

3

Helix

54 – 63

10

Helix

67 – 69

3

Beta strand

71 – 76

6

Helix

78 – 80

3

Helix

86 – 97

12

Beta strand

102 – 107

6

Helix

112 – 114

3

Helix

117 – 129

13

Beta strand

132 – 140

9

Helix

143 – 153

11

Beta strand

158 – 163

6

Helix

171 – 179

9

Beta strand

183 – 188

6

Helix

191 – 193

3

Helix

201 – 210

10

Helix

214 – 224

11

Helix

236 – 243

8

Helix

252 – 259

8

Beta strand

264 – 266

3

Turn

274 – 276

3

Sequences

	Sequence		Length	Mass (Da)
	P06632 [UniParc]. Last modified January 23, 2007. Version 3. Checksum: 403A2E9AA4624A37	FASTA	278	30,119
	10 20 30 40 50 60 MTVPSIVLND GNSIPQLGYG VFKVPPADTQ RAVEEALEVG YRHIDTAAIY GNEEGVGAAI 70 80 90 100 110 120 AASGIARDDL FITTKLWNDR HDGDEPAAAI AESLAKLALD QVDLYLVHWP TPAADNYVHA 130 140 150 160 170 180 WEKMIELRAA GLTRSIGVSN HLVPHLERIV AATGVVPAVN QIELHPAYQQ REITDWAAAH 190 200 210 220 230 240 DVKIESWGPL GQGKYDLFGA EPVTAAAAAH GKTPAQAVLR WHLQKGFVVF PKSVRRERLE 250 260 270 ENLDVFDFDL TDTEIAAIDA MDPGDGSGRV SAHPDEVD

References

[1]	"Production of 2-keto-L-gulonate, an intermediate in L-ascorbate synthesis, by a genetically modified Erwinia herbicola." Anderson S., Marks C.B., Lazarus R.A., Miller J.V., Stafford K., Seymour J., Light D., Rastetter W., Estell D.A. Science 230:144-149(1985) Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA], PROTEIN SEQUENCE OF 2-41; 76-89; 97-123 AND 184-194, CHARACTERIZATION.
[2]	"Purification and characterization of 2,5-diketo-D-gluconate reductase from Corynebacterium sp." Miller J.V., Estell D.A., Lazarus R.A. J. Biol. Chem. 262:9016-9020(1987) [PubMed] [Europe PMC] [Abstract] Cited for: PROTEIN SEQUENCE OF 2-41, CHARACTERIZATION, BIOPHYSICOCHEMICAL PROPERTIES.
[3]	"Molecular modeling of substrate binding in wild-type and mutant Corynebacteria 2,5-diketo-D-gluconate reductases." Khurana S., Sanli G., Powers D.B., Anderson S., Blaber M. Proteins 39:68-75(2000) [PubMed] [Europe PMC] [Abstract] Cited for: 3D-STRUCTURE MODELING OF SUBSTRATE-BINDING SITE.
[4]	"Crystal structure of 2,5-diketo-D-gluconic acid reductase A complexed with NADPH at 2.1-A resolution." Khurana S., Powers D.B., Anderson S., Blaber M. Proc. Natl. Acad. Sci. U.S.A. 95:6768-6773(1998) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.1 ANGSTROMS).
[5]	"Structural assembly of the active site in an aldo-keto reductase by NADPH cofactor." Sanli G., Blaber M. J. Mol. Biol. 309:1209-1218(2001) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (1.9 ANGSTROMS).
[6]	"Structural alteration of cofactor specificity in Corynebacterium 2,5-diketo-D-gluconic acid reductase." Sanli G., Banta S., Anderson S., Blaber M. Protein Sci. 13:504-512(2004) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.0 ANGSTROMS) OF MUTANT WITH TYR-22; GLY-232; HIS-238 AND GLY-272 IN COMPLEX WITH NADH.

Cross-references

Sequence databases

EMBL
GenBank
DDBJ

M12799 Genomic DNA. Translation: AAA83534.1.

PIR

I40838.

3D structure databases

PDBe
RCSB PDB
PDBj

Entry	Method	Resolution (Å)	Chain	Positions	PDBsum
1A80	X-ray	2.10	A	2-278	[»]
1HW6	X-ray	1.90	A	1-278	[»]
1M9H	X-ray	2.00	A	1-278	[»]

ProteinModelPortal

P06632.

SMR

P06632. Positions 2-278.

ModBase

Search...

Protocols and materials databases

StructuralBiologyKnowledgebase

Search...

Enzyme and pathway databases

BioCyc

MetaCyc:MONOMER-17833.

Family and domain databases

Gene3D

3.20.20.100. 1 hit.

InterPro

IPR001395. Aldo/ket_red.
IPR018170. Aldo/ket_reductase_CS.
IPR020471. Aldo/keto_reductase_subgr.
IPR023210. NADP_OxRdtase_dom.
[Graphical view]

PANTHER

PTHR11732. PTHR11732. 1 hit.

Pfam

PF00248. Aldo_ket_red. 1 hit.
[Graphical view]

PIRSF

PIRSF000097. AKR. 1 hit.

PRINTS

PR00069. ALDKETRDTASE.

SUPFAM

SSF51430. Aldo/ket_red. 1 hit.

PROSITE

PS00798. ALDOKETO_REDUCTASE_1. 1 hit.
PS00062. ALDOKETO_REDUCTASE_2. False negative.
PS00063. ALDOKETO_REDUCTASE_3. False negative.
[Graphical view]

ProtoNet

Search...

Other

EvolutionaryTrace

P06632.

Entry information

Entry name

DKGA_CORSC

Accession

Primary (citable) accession number: P06632

Entry history

Integrated into UniProtKB/Swiss-Prot:	November 1, 1988
Last sequence update:	January 23, 2007
Last modified:	May 29, 2013
This is version 80 of the entry and version 3 of the sequence. [Complete history]

Entry status

Reviewed (UniProtKB/Swiss-Prot)

Annotation program

Prokaryotic Protein Annotation Program