Q8GCH2 (GUMK2_XANCP) Reviewed, UniProtKB/Swiss-Prot

Names and origin

Protein names

Recommended name:
UDP-glucuronate:glycolipid 2-beta-glucuronosyltransferase
Short name=UDP-GlcA:glycolipid glucuronosyltransferase
EC=2.4.1.264

Alternative name(s):
D-man-alpha-(1->3)-D-Glc-beta-(1->4)-D-Glc-alpha-1-diphosphoundecaprenol 2-beta-glucuronyltransferase

Gene names

Name:

gumK

Organism

Xanthomonas campestris pv. campestris

Taxonomic identifier

340 [NCBI]

Taxonomic lineage

Bacteria › Proteobacteria › Gammaproteobacteria › Xanthomonadales › Xanthomonadaceae › Xanthomonas

Protein attributes

Sequence length	400 AA.
Sequence status	Complete.
Protein existence	Evidence at protein level

General annotation (Comments)

Function	Catalyzes the transfer of a glucuronic acid (GlcA) residue from UDP-glucuronate to mannose-alpha-1,3-glucose-beta-1,4-glucose-P-P-polyisoprenyl to form the lipid-linked tetrasaccharide GlcA-Man-Glc(2)-PP-Pol, with a glucuronic acid-beta-mannose linkage. Is involved in the biosynthesis of the exopolysaccharide xanthan, since it catalyzes the fourth glycosylation step in the assembly of the pentasaccharide-P-P-polyisoprenyl repeating unit of xanthan. Is unable to use the trisaccharide acceptor freed from the pyrophosphate lipid moiety. Does not show specificity for the lipidic portion of the acceptor. Shows diminished activity when tested with 6-O-acetyl-mannose-alpha-1,3-glucose-beta-1,4-glucose-P-P-polyisoprenyl, a putative intermediate in the synthesis of xanthan; this could indicate that acetylation of the internal mannose takes place after the formation of the GumK product. Ref.2
Catalytic activity	UDP-glucuronate + D-Man-alpha-(1->3)-D-Glc-beta-(1->4)-D-Glc-alpha-1-diphospho-ditrans,octacis-undecaprenol = UDP + D-GlcA-beta-(1->2)-D-Man-alpha-(1->3)-D-Glc-beta-(1->4)-D-Glc-alpha-1-diphospho-ditrans,octacis-undecaprenol. Ref.2 Ref.5
Pathway	Glycan biosynthesis; xanthan biosynthesis. Ref.2
Subcellular location	Cell inner membrane; Peripheral membrane protein; Cytoplasmic side Ref.2.
Disruption phenotype	Cells lacking this gene accumulate Man-alpha-1,3-Glc-beta-1,4-Glc-PP-polyisoprenyl, are unable to produce GlcA-Man-Glc(2)-PP-Pol, and show a decrease in xanthan production of more than 95%. Ref.2 Ref.3
Sequence similarities	Belongs to the glycosyltransferase 70 family.
Biophysicochemical properties	Kinetic parameters: KM=62 µM for UDP-glucuronate Ref.5 KM=198 µM for Man-Glc(2)-PP-Pol Vmax=116 pmol/min/µg enzyme
Sequence caution	The sequence AAA86379.1 differs from that shown. Reason: Erroneous initiation. Translation N-terminally extended.

Ontologies

Keywords
Biological process	Carbohydrate metabolism
Cellular component	Cell inner membrane Cell membrane Membrane
Molecular function	Glycosyltransferase Transferase
Technical term	3D-structure
Gene Ontology (GO)
Biological process	polysaccharide biosynthetic process Inferred from mutant phenotype Ref.2. Source: UniProtKB
Cellular component	plasma membrane Inferred from direct assay Ref.2. Source: UniProtKB
Molecular function	glucuronosyltransferase activity Inferred from direct assay Ref.2. Source: UniProtKB
Complete GO annotation...

Sequence annotation (Features)

Feature key

Position(s)

Length

Description

Graphical view

Feature identifier

Molecule processing

Chain

1 – 400

400

UDP-glucuronate:glycolipid 2-beta-glucuronosyltransferase

PRO_0000414019

Regions

Region

230 – 231

2

UDP-glucuronate binding

Region

272 – 273

2

UDP-glucuronate binding

Region

306 – 310

5

UDP-glucuronate binding

Sites

Active site

157

1

Proton acceptor Ref.5

Binding site

292

1

UDP-glucuronate

Experimental info

Mutagenesis

157

1

D → A, E or N: Loss of catalytic activity. Loss of xanthan production. Ref.5

Mutagenesis

192

1

E → A: No effect on both substrate affinity and catalytic activity. Ref.5

Mutagenesis

207

1

D → A: No effect on both substrate affinity and catalytic activity. Ref.5

Mutagenesis

231

1

M → A: No effect on both substrate affinity and catalytic activity. Ref.5

Mutagenesis

234

1

D → A: No effect on both substrate affinity and catalytic activity. Ref.5

Mutagenesis

272

1

E → A: 2-fold decrease in both affinity for UDP-GlcA and catalytic activity. Ref.5

Mutagenesis

292

1

Y → A: 14-fold decrease in catalytic efficiency. 25% of wild-type xanthan production. Ref.5

Mutagenesis

307

1

K → A: 54-fold decrease in catalytic efficiency. 30% of wild-type xanthan production. Ref.5

Mutagenesis

310

1

Q → A: 19-fold decrease in affinity for UDP-GlcA but no effect on catalytic activity. 60% of wild-type xanthan production. Ref.5

Sequence conflict

2

1

S → G in AAN86640. Ref.2

Sequence conflict

2

1

S → G in AAA86379. Ref.2

Sequence conflict

22

1

A → S in AAN86640. Ref.2

Sequence conflict

22

1

A → S in AAA86379. Ref.2

Sequence conflict

194

1

A → V in AAN86640. Ref.2

Sequence conflict

194

1

A → V in AAA86379. Ref.2

Secondary structure

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400

Helix Strand Turn

Details...

Beta strand

17 – 23

7

Beta strand

27 – 29

3

Helix

32 – 43

12

Beta strand

46 – 50

5

Helix

57 – 60

4

Helix

65 – 70

6

Beta strand

73 – 77

5

Beta strand

80 – 84

5

Beta strand

87 – 89

3

Helix

97 – 99

3

Helix

100 – 112

13

Helix

116 – 124

9

Beta strand

126 – 133

8

Helix

134 – 138

5

Helix

139 – 145

7

Beta strand

149 – 157

9

Helix

159 – 162

4

Helix

166 – 175

10

Helix

176 – 179

4

Beta strand

180 – 185

6

Helix

187 – 192

6

Beta strand

199 – 201

3

Helix

210 – 214

5

Beta strand

221 – 228

8

Helix

235 – 244

10

Beta strand

248 – 254

7

Beta strand

267 – 270

4

Helix

275 – 283

9

Beta strand

286 – 289

4

Helix

301 – 304

4

Helix

306 – 313

8

Beta strand

318 – 321

4

Helix

322 – 324

3

Beta strand

329 – 334

6

Helix

339 – 351

13

Helix

363 – 371

9

Helix

373 – 375

3

Helix

377 – 379

3

Beta strand

380 – 382

3

Sequences

	Sequence		Length	Mass (Da)
	Q8GCH2 [UniParc]. Last modified November 16, 2011. Version 2. Checksum: 250231946AB7EB61	FASTA	400	44,438
	10 20 30 40 50 60 MSVSPAAPAS GIRRPCYLVL SAHDFRTPRR ANIHFITDQL ALRGTTRFFS LRYSRLSRMK 70 80 90 100 110 120 GDMRLPLDDT ANTVVSHNGV DCYLWRTTVH PFNTRRSWLR PVEDAMFRWY AAHPPKQLLD 130 140 150 160 170 180 WMRESDVIVF ESGIAVAFIE LAKRVNPAAK LVYRASDGLS TINVASYIER EFDRVAPTLD 190 200 210 220 230 240 VIALVSPAMA AEVASRDNVF HVGHGVDHNL DQLGDPSPYA EGIHAVAVGS MLFDPEFFVV 250 260 270 280 290 300 ASKAFPQVTF HVIGSGMGRH PGYGDNVIVY GEMKHAQTIG YIKHARFGIA PYASEQVPVY 310 320 330 340 350 360 LADSSMKLLQ YDFFGLPAVC PNAVVGPYKS RFGYTPGNAD SVIAAITQAL EAPRVRYRQC 370 380 390 400 LNWSDTTDRV LDPRAYPETR LYPHPPTAAP QLSSEAALSH

References

[1]	"Recombinant-DNA mediated production of xanthan gum." Capage M.A., Doherty D.H., Betlach M.R., Vanderslice R.W. Submitted (APR-1995) to the EMBL/GenBank/DDBJ databases Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA]. Strain: ATCC 13951 / NCIB 11803 / NRRL B-1459.
[2]	"Functional characterization of GumK, a membrane-associated beta-glucuronosyltransferase from Xanthomonas campestris required for xanthan polysaccharide synthesis." Barreras M., Abdian P.L., Ielpi L. Glycobiology 14:233-241(2004) [PubMed: 14736729] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA], IDENTIFICATION OF START SITE, FUNCTION, CATALYTIC ACTIVITY, ROLE IN XANTHAN BIOSYNTHESIS, SUBSTRATE SPECIFICITY, SUBCELLULAR LOCATION, DISRUPTION PHENOTYPE, PATHWAY. Strain: ATCC 13951 / NCIB 11803 / NRRL B-1459.
[3]	"Xanthomonas campestris pv. campestris gum mutants: effects on xanthan biosynthesis and plant virulence." Katzen F., Ferreiro D.U., Oddo C.G., Ielmini M.V., Becker A., Puhler A., Ielpi L. J. Bacteriol. 180:1607-1617(1998) [PubMed: 9537354] [Abstract] Cited for: DISRUPTION PHENOTYPE. Strain: ATCC 13951 / NCIB 11803 / NRRL B-1459.
[4]	"Crystallization and preliminary crystallographic characterization of GumK, a membrane-associated glucuronosyltransferase from Xanthomonas campestris required for xanthan polysaccharide synthesis." Barreras M., Bianchet M.A., Ielpi L. Acta Crystallogr. F 62:880-883(2006) [PubMed: 16946469] [Abstract] Cited for: CRYSTALLIZATION. Strain: ATCC 13951 / NCIB 11803 / NRRL B-1459.
[5]	"Structure and mechanism of GumK, a membrane-associated glucuronosyltransferase." Barreras M., Salinas S.R., Abdian P.L., Kampel M.A., Ielpi L. J. Biol. Chem. 283:25027-25035(2008) [PubMed: 18596046] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (1.90 ANGSTROMS) OF WILD-TYPE AND MUTANT ALA-157 APOENZYME AND IN COMPLEX WITH UDP, CATALYTIC ACTIVITY, KINETIC PARAMETERS, MUTAGENESIS OF ASP-157; GLU-192; ASP-207; MET-231; ASP-234; GLU-272; TYR-292; LYS-307 AND GLN-310, ACTIVE SITE, CATALYTIC MECHANISM. Strain: ATCC 13951 / NCIB 11803 / NRRL B-1459.

Cross-references

Sequence databases

EMBL
GenBank
DDBJ

U22511 Genomic DNA. Translation: AAA86379.1. Different initiation.
AY170889 Genomic DNA. Translation: AAN86640.1.

PIR

S67860.

3D structure databases

PDBe
RCSB PDB
PDBj

Entry	Method	Resolution (Å)	Chain	Positions	PDBsum
2HY7	X-ray	1.90	A	1-400	[»]
2Q6V	X-ray	2.28	A	1-400	[»]
3CUY	X-ray	2.30	A	1-400	[»]
3CV3	X-ray	2.25	A	1-400	[»]

ProteinModelPortal

Q8GCH2.

ModBase

Search...

Protein family/group databases

CAZy

GT70. Glycosyltransferase Family 70.

Protocols and materials databases

StructuralBiologyKnowledgebase

Search...

Family and domain databases

ProtoNet

Search...

Entry information

Entry name

GUMK2_XANCP

Accession

Primary (citable) accession number: Q8GCH2
Secondary accession number(s): Q56777

Entry history

Integrated into UniProtKB/Swiss-Prot:	November 16, 2011
Last sequence update:	November 16, 2011
Last modified:	January 25, 2012
This is version 22 of the entry and version 2 of the sequence. [Complete history]

Entry status

Reviewed (UniProtKB/Swiss-Prot)

Annotation program

Prokaryotic Protein Annotation Program