Reviewed, UniProtKB/Swiss-Prot Q9ZEU2 (AMYS_NEIPO)

Names and origin

Protein names

Recommended name:
    Amylosucrase
    EC=2.4.1.4

Gene names

Name:

ams

Organism

Neisseria polysaccharea

Taxonomic identifier

489 [NCBI]

Taxonomic lineage

Bacteria › Proteobacteria › Betaproteobacteria › Neisseriales › Neisseriaceae › Neisseria

Protein attributes

Sequence length	636 AA.
Sequence status	Complete.
Sequence processing	The displayed sequence is not processed.
Protein existence	Evidence at protein level.

General annotation (Comments)

Function	Catalyzes the synthesis of alpha-glucan from sucrose. Catalyzes, in addition, sucrose hydrolysis, maltose and maltotriose synthesis by successive transfers of the glucosyl moiety of sucrose onto the released glucose, and finally turanose and trehalulose synthesis, these two sucrose isomers being obtained by glucosyl transfer onto fructose. Ref.2
Catalytic activity	Sucrose + ((1->4)-alpha-D-glucosyl)(n) = D-fructose + ((1->4)-alpha-D-glucosyl)(n+1).
Enzyme regulation	Amylosucrase favors hydrolysis at low sucrose concentrations, and polymerization at high sucrose concentrations. Competitively inhibited by fructose.
Subunit structure	Monomer. Ref.2
Subcellular location	Secreted. Ref.1
Miscellaneous	Like the recombinant protein expressed in E.coli, the amylosucrase may be secreted in N.polysaccharea without cleavage of a signal sequence.
Sequence similarities	Belongs to the glycosyl hydrolase 13 family.
Biophysicochemical properties	Kinetic parameters: This enzyme does not present a classic Michaelis-Menten behavior for sucrose consumption, that could be related to the presence of a second sucrose binding site. Nevertheless, it is possible to model sucrose consumption rate versus sucrose concentration by two different Michaelis-Menten equations whose apparent kinetic constants are given just above. KM=1.9 mM for sucrose (in the sucrose consumption assay, when initial sucrose < 20 mM) Ref.4 KM=1.7 mM for sucrose (in the sucrose hydrolysis reaction, when initial sucrose < 20 mM) KM=1.9 mM for sucrose (in the polymerization reaction, when initial sucrose < 20 mM) KM=50.2 mM for sucrose (in the sucrose consumption assay, when initial sucrose > 20 mM) KM=38.7 mM for sucrose (in the sucrose hydrolysis reaction, when initial sucrose > 20 mM) Vmax=470 µmol/min/g enzyme for sucrose consumption (when initial sucrose < 20 mM) Vmax=288 µmol/min/g enzyme for sucrose hydrolysis (when initial sucrose < 20 mM) Vmax=147 µmol/min/g enzyme for polymerization reaction (when initial sucrose < 20 mM) Vmax=1100 µmol/min/g enzyme for sucrose consumption (when initial sucrose > 20 mM) Vmax=472 µmol/min/g enzyme for sucrose hydrolysis (when initial sucrose > 20 mM) Vmax=1620 µmol/min/g enzyme for polymerization reaction (when initial sucrose > 20 mM)
Sequence caution	The sequence described in Ref.1 differs from that shown. Reason: Frameshift at positions 183, 203, 274, 313, 507, 538 and 579.

Ontologies

Keywords
Cellular component	Secreted
Molecular function	Glycosyltransferase Transferase
Technical term	3D-structure Direct protein sequencing
Gene Ontology (GO)
Biological process	carbohydrate metabolic process Inferred from electronic annotation. Source: InterPro
Cellular component	extracellular region Inferred from electronic annotation. Source: UniProtKB-SubCell
Molecular function	amylosucrase activity Inferred from electronic annotation. Source: EC cation binding 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 – 636

636

Amylosucrase

PRO_0000045154

Sites

Active site

294

1

Nucleophile

Active site

336

1

Proton donor

Binding site

152

1

Substrate

Binding site

195

1

Substrate

Binding site

262

1

Substrate

Binding site

292

1

Substrate

Binding site

400

1

Substrate

Binding site

401

1

Substrate

Binding site

517

1

Substrate

Experimental info

Mutagenesis

195

1

H → Q: 98% reduction of activity. Ref.5

Mutagenesis

294

1

D → E or N: Complete loss of activity. Ref.5

Mutagenesis

308

1

E → Q: No effect. Ref.5

Mutagenesis

336

1

E → Q: Complete loss of activity. Ref.5

Mutagenesis

352

1

E → Q: 30% reduction of activity. Ref.5

Mutagenesis

400

1

H → N: 97% reduction of activity. Ref.5

Mutagenesis

401

1

D → E: Almost complete loss of activity. Ref.5

Sequence conflict

14

1

Y → H in AAM51153. Ref.3

Sequence conflict

84 – 85

2

NS → SA in AAM51153. Ref.3

Sequence conflict

89

1

D → N in AAM51153. Ref.3

Sequence conflict

93

1

A → E in AAM51153. Ref.3

Sequence conflict

159

1

S → T AA sequence Ref.1

Sequence conflict

171

1

G → D in AAM51153. Ref.3

Sequence conflict

208

1

Missing AA sequence Ref.1

Sequence conflict

392

1

A → S in AAM51153. Ref.3

Sequence conflict

437

1

S → T AA sequence Ref.1

Sequence conflict

449

1

S → N in AAM51153. Ref.3

Sequence conflict

459

1

A → S in AAM51153. Ref.3

Sequence conflict

463

1

V → A in AAM51153. Ref.3

Sequence conflict

468

1

D → N in AAM51153. Ref.3

Sequence conflict

474

1

D → S in AAM51153. Ref.3

Sequence conflict

501

1

L → P in AAM51153. Ref.3

Sequence conflict

505 – 510

6

DWSQDS → GWAQDG in AAM51153. Ref.3

Secondary structure

1

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636

Helix Strand Turn

Details...

Helix

13 – 19

7

Helix

20 – 22

3

Helix

25 – 33

9

Helix

35 – 59

25

Helix

65 – 82

18

Helix

85 – 95

11

Helix

98 – 102

5

Beta strand

108 – 111

4

Helix

113 – 117

5

Helix

120 – 125

6

Helix

127 – 133

7

Beta strand

136 – 140

5

Turn

152 – 155

4

Beta strand

160 – 163

4

Turn

165 – 167

3

Helix

170 – 182

13

Beta strand

186 – 191

6

Beta strand

194 – 197

4

Helix

201 – 207

7

Helix

211 – 213

3

Beta strand

219 – 223

5

Helix

224 – 229

6

Turn

230 – 232

3

Turn

238 – 240

3

Beta strand

245 – 247

3

Beta strand

253 – 255

3

Beta strand

257 – 259

3

Beta strand

262 – 265

4

Helix

270 – 284

15

Turn

285 – 287

3

Beta strand

289 – 293

5

Helix

296 – 298

3

Beta strand

307 – 309

3

Helix

311 – 327

17

Beta strand

332 – 335

4

Helix

341 – 344

4

Helix

345 – 347

3

Beta strand

352 – 357

6

Helix

359 – 371

13

Helix

375 – 383

9

Beta strand

392 – 397

6

Helix

409 – 414

6

Helix

419 – 430

12

Beta strand

442 – 444

3

Turn

448 – 450

3

Beta strand

454 – 456

3

Helix

459 – 463

5

Helix

465 – 467

3

Helix

472 – 485

14

Beta strand

486 – 493

8

Helix

496 – 498

3

Helix

506 – 508

3

Turn

510 – 514

5

Helix

516 – 520

5

Helix

526 – 529

4

Turn

530 – 533

4

Helix

538 – 555

18

Helix

557 – 559

3

Beta strand

574 – 579

6

Turn

580 – 582

3

Beta strand

583 – 588

6

Beta strand

590 – 592

3

Beta strand

594 – 596

3

Turn

598 – 603

6

Beta strand

606 – 610

5

Turn

611 – 613

3

Beta strand

616 – 618

3

Beta strand

623 – 625

3

Beta strand

630 – 634

5

Sequences

	Sequence		Length	Mass (Da)
	Q9ZEU2-1 [UniParc]. Last modified May 1, 1999. Version 1. Checksum: B7656C19BF1A2065	FASTA	636	72,344
	10 20 30 40 50 60 MLTPTQQVGL ILQYLKTRIL DIYTPEQRAG IEKSEDWRQF SRRMDTHFPK LMNELDSVYG 70 80 90 100 110 120 NNEALLPMLE MLLAQAWQSY SQRNSSLKDI DIARENNPDW ILSNKQVGGV CYVDLFAGDL 130 140 150 160 170 180 KGLKDKIPYF QELGLTYLHL MPLFKCPEGK SDGGYAVSSY RDVNPALGTI GDLREVIAAL 190 200 210 220 230 240 HEAGISAVVD FIFNHTSNEH EWAQRCAAGD PLFDNFYYIF PDRRMPDQYD RTLREIFPDQ 250 260 270 280 290 300 HPGGFSQLED GRWVWTTFNS FQWDLNYSNP WVFRAMAGEM LFLANLGVDI LRMDAVAFIW 310 320 330 340 350 360 KQMGTSCENL PQAHALIRAF NAVMRIAAPA VFFKSEAIVH PDQVVQYIGQ DECQIGYNPL 370 380 390 400 410 420 QMALLWNTLA TREVNLLHQA LTYRHNLPEH TAWVNYVRSH DDIGWTFADE DAAYLGISGY 430 440 450 460 470 480 DHRQFLNRFF VNRFDGSFAR GVPFQYNPST GDCRVSGTAA ALVGLAQDDP HAVDRIKLLY 490 500 510 520 530 540 SIALSTGGLP LIYLGDEVGT LNDDDWSQDS NKSDDSRWAH RPRYNEALYA QRNDPSTAAG 550 560 570 580 590 600 QIYQGLRHMI AVRQSNPRFD GGRLVTFNTN NKHIIGYIRN NALLAFGNFS EYPQTVTAHT 610 620 630 LQAMPFKAHD LIGGKTVSLN QDLTLQPYQV MWLEIA

References

[1]	"Cloning and characterization of the gene for amylosucrase from Neisseria polysaccharea: production of a linear alpha-1,4-glucan." Buettcher V., Welsh T., Willmitzer L., Kossmann J. J. Bacteriol. 179:3324-3330(1997) [PubMed: 9150231] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA], PROTEIN SEQUENCE OF N-TERMINUS, SUBCELLULAR LOCATION. Strain: ATCC 43768 / CCUG 18030 / CIP 100113 / NCTC 11858.
[2]	"Sequence analysis of the gene encoding amylosucrase from Neisseria polysaccharea and characterization of the recombinant enzyme." Potocki de Montalk G., Remaud-Simeon M., Willemot R.-M., Planchot V., Monsan P. J. Bacteriol. 181:375-381(1999) [PubMed: 9882648] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA], FUNCTION, SUBUNIT. Strain: ATCC 43768 / CCUG 18030 / CIP 100113 / NCTC 11858.
[3]	"Nonencapsulated Neisseria meningitidis strain produces amylopectin from sucrose: altering the concept for differentiation between N. meningitidis and N. polysaccharea." Zhu P., Tsang R.S.W., Tsai C.-M. J. Clin. Microbiol. 41:273-278(2003) [PubMed: 12517860] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA]. Strain: 85322.
[4]	"Amylosucrase from Neisseria polysaccharea: novel catalytic properties." Potocki de Montalk G., Remaud-Simeon M., Willemot R.-M., Sarcabal P., Planchot V., Monsan P. FEBS Lett. 471:219-223(2000) [PubMed: 10767427] [Abstract] Cited for: CHARACTERIZATION, KINETIC PARAMETERS. Strain: ATCC 43768 / CCUG 18030 / CIP 100113 / NCTC 11858.
[5]	"Identification of key amino acid residues in Neisseria polysaccharea amylosucrase." Sarcabal P., Remaud-Simeon M., Willemot R.-M., Potocki de Montalk G., Svensson B., Monsan P. FEBS Lett. 474:33-37(2000) [PubMed: 10828446] [Abstract] Cited for: MUTAGENESIS OF HIS-195; ASP-294; GLU-308; GLU-336; GLU-352; HIS-400 AND ASP-401. Strain: ATCC 43768 / CCUG 18030 / CIP 100113 / NCTC 11858.
[6]	"Crystal structures of amylosucrase from Neisseria polysaccharea in complex with D-glucose and the active site mutant Glu328Gln in complex with the natural substrate sucrose." Mirza O., Skov L.K., Remaud-Simeon M., Potocki de Montalk G., Albenne C., Monsan P., Gajhede M. Biochemistry 40:9032-9039(2001) [PubMed: 11467966] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (1.66 ANGSTROMS) OF 13-636 IN COMPLEX WITH D-GLUCOSE. Strain: ATCC 43768 / CCUG 18030 / CIP 100113 / NCTC 11858.
[7]	"Amylosucrase, a glucan-synthesizing enzyme from the alpha-amylase family." Skov L.K., Mirza O., Henriksen A., De Montalk G.P., Remaud-Simeon M., Sarcabal P., Willemot R.M., Monsan P., Gajhede M. J. Biol. Chem. 276:25273-25278(2001) [PubMed: 11306569] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (1.4 ANGSTROMS) OF 13-636. Strain: ATCC 43768 / CCUG 18030 / CIP 100113 / NCTC 11858.
[8]	"Oligosaccharide and sucrose complexes of amylosucrase. Structural implications for the polymerase activity." Skov L.K., Mirza O., Sprogoe D., Dar I., Remaud-Simeon M., Albenne C., Monsan P., Gajhede M. J. Biol. Chem. 277:47741-47747(2002) [PubMed: 12364331] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.2 ANGSTROMS) OF 13-636 OF MUTANT GLN-336 IN COMPLEX WITH MALTOHEPTAOSE.
[9]	"Crystal structure of the covalent intermediate of amylosucrase from Neisseria polysaccharea." Jensen M.H., Mirza O., Albenne C., Remaud-Simeon M., Monsan P., Gajhede M., Skov L.K. Biochemistry 43:3104-3110(2004) [PubMed: 15023061] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.2 ANGSTROMS) OF 13-636 OF MUTANT GLN-336 IN COMPLEX WITH ALPHA-D-GLUCOPYRANOSYL FLUORIDE.

Cross-references

Sequence databases

EMBLGenBankDDBJ

AJ011781 Genomic DNA. Translation: CAA09772.1.
AY099335 Genomic DNA. Translation: AAM51153.1.

3D structure databases

PDBPDBe

Entry	Method	Resolution (Å)	Chain	Positions	PDBsum
1G5A	X-ray	1.40	A	13-636	[»]
1JG9	X-ray	1.66	A	13-636	[»]
1JGI	X-ray	2.00	A	13-636	[»]
1MVY	X-ray	2.00	A	13-636	[»]
1MW0	X-ray	2.01	A	13-636	[»]
1MW1	X-ray	2.10	A	13-636	[»]
1MW2	X-ray	2.10	A	13-636	[»]
1MW3	X-ray	2.00	A	13-636	[»]
1S46	X-ray	2.20	A	13-636	[»]
1ZS2	X-ray	2.16	A	13-636	[»]

ModBase

Search...

Protein family/group databases

CAZy

GH13. Glycoside Hydrolase Family 13.

Enzyme and pathway databases

BRENDA

2.4.1.4. 264966.

Family and domain databases

InterPro

IPR006047. Glyco_hydro_13_cat.
IPR013781. Glyco_hydro_sg_catalytic.
[Graphical view]

Gene3D

G3DSA:3.20.20.80. Glyco_hydro_cat. 1 hit.

Pfam

PF00128. Alpha-amylase. 1 hit.
[Graphical view]

ProtoNet

Search...

Entry information

Entry name

AMYS_NEIPO

Accession

Primary (citable) accession number: Q9ZEU2
Secondary accession number(s): Q84HD5

Entry history

Integrated into UniProtKB/Swiss-Prot:	December 20, 2005
Last sequence update:	May 1, 1999
Last modified:	June 16, 2009
This is version 45 of the entry and version 1 of the sequence. [Complete history]

Entry status

Reviewed (UniProtKB/Swiss-Prot)

Annotation project

HAMAP (High-quality Automated and Manual Annotation of microbial Proteomes)