Information on EC 2.4.1.25 - 4-alpha-glucanotransferase and Organism(s) Corynebacterium glutamicum and UniProt Accession Q8NNA7

-

-

?

maltotriose + glycosyl acceptor

maltose + D-glucose + maltooligosaccharides

the wild type enzyme prefers maltotriose for disproportionation reaction

-

?

maltotriose + maltotriose

maltooligosaccharides

-

-

?

pea starch + glycosyl acceptor

cycloamylose + ?

-

-

?

pea starch + glycosyl acceptor

large-ring cyclodextrins

-

-

?

potato starch + glycosyl acceptor

?

-

-

?

cassava starch + glycosyl acceptor

?

-

-

?

corn starch + glycosyl acceptor

large-ring cyclodextrins

-

-

?

glutinous-rice starch + glycosyl acceptor

large-ring cyclodextrins

-

-

?

maltoheptaose + glycosyl acceptor

maltose + D-glucose + maltooligosaccharides

-

worst substrate

-

?

maltoheptaose + maltoheptaose

?

-

-

?

maltohexaose + glycosyl acceptor

maltose + D-glucose + maltooligosaccharides

-

-

?

maltohexaose + maltohexaose

maltooligosaccharide

-

-

?

maltopentaose + glycosyl acceptor

maltose + D-glucose + maltooligosaccharides

-

-

?

maltopentaose + maltopentaose

maltooligosaccharides

-

-

?

maltose + glycosyl acceptor

?

-

lowest activity

-

?

maltose + maltose

?

-

worst substrate

-

?

maltotetraose + glycosyl acceptor

maltose + D-glucose + maltooligosaccharides

-

second best substrate

-

?

maltotetraose + maltotetraose

maltooligosaccharides

-

-

?

maltotriose

maltooligosaccharides

-

-

?

maltotriose + glycosyl acceptor

maltose + D-glucose + maltooligosaccharides

-

best substrate

-

?

maltotriose + maltotriose

maltooligosaccharides

-

most efficient substrate

-

?

pea starch + glycosyl acceptor

?

-

-

?

pea starch + glycosyl acceptor

large ring-cyclodextrins 23-26

-

-

?

pea starch + glycosyl acceptor

large ring-cyclodextrins 29-33

-

-

?

pea starch + glycosyl acceptor

large-ring cyclodextrins

-

-

?

potato starch + maltose

large-ring cyclodextrins

-

-

?

rice starch + glycosyl acceptor

large-ring cyclodextrins

-

-

?

soluble potato starch + glycosyl acceptor

?

-

-

?

soluble potato starch + maltose

?

-

-

?

tapioca starch + glycosyl acceptor

large-ring cyclodextrins

-

-

?

N-bromosuccinimide

51.2% residual activity at 0.1 mM

16.9 - 37.4

maltotriose

6 entries

15.4 - 18.6

maltopentaose

12.9 - 20.4

maltotriose

11 entries

120.8 - 3033

maltotriose

6 entries

0.025 - 4.8

maltopentaose

0.033 - 2800

maltotriose

11 entries

3.2 - 180

maltotriose

6 entries

0.0017 - 0.26

maltopentaose

0.0023 - 163.3

maltotriose

11 entries

21.8

-

wild-type, pH 6.0, 30°C

6.2

-

mutant Y172A, pH 6.0, 30°C

6

-

718654, 736255, 755942

5.5 - 9

-

30

45

-

735379, 736249, 755897, 757031

UniProt

81000

* 81000, SDS-PAGE

81000

-

x * 81000, SDS-PAGE

?

monomer or dimer

the enzyme forms a dimer without NaCl and exists as a monomer in physiological concentration of NaCl

?

hanging drop vapor diffusion method, using 2.0 M ammonium sulfate and 0.1 M bis-tris methane (pH 6.5)

sitting drop vapor diffusion method, using 0.1 M bis-Tris pH 5.5 and 2.0 M ammonium sulfate

H461A

the mutation leads to a significant (8.6fold) decrease in transglucosylation activity compared to the wild type enzyme, while hydrolysis activity is barely affected. The mutant cannot produce large-ring cyclodextrins from maltotriose and prefers maltose over maltotriose as substrate

H461D

the mutation leads to a significant (3.4fold) decrease in transglucosylation activity compared to the wild type enzyme, while hydrolysis activity is barely affected. The mutant cannot produce large-ring cyclodextrins from maltotriose and prefers maltose over maltotriose as substrate

H461R

the mutation leads to a significant (6fold) decrease in transglucosylation activity compared to the wild type enzyme, while hydrolysis activity is barely affected. The mutant cannot produce large-ring cyclodextrins from maltotriose and prefers maltose over maltotriose as substrate

H461S

the mutation leads to a significant (3.4fold) decrease in transglucosylation activity compared to the wild type enzyme, while hydrolysis activity is barely affected. The mutant cannot produce large-ring cyclodextrins from maltotriose and prefers maltose over maltotriose as substrate

H461W

the mutation leads to a significant (6fold) decrease in transglucosylation activity compared to the wild type enzyme, while hydrolysis activity is barely affected. The mutant cannot produce large-ring cyclodextrins from maltotriose and prefers maltose over maltotriose as substrate

W330A

inactive

W425A

inactive

W673A

inactive

A406L

-

the mutant shows higher thermostability at 35-40°C, higher intermolecular transglucosylation activity with an upward shift in the optimum temperature and a slight increase in the optimum pH for disproportionation and cyclization reactions compared to the wild type enzyme. The mutant shows higher specific activities for starch transglucosylation (2.1fold) and disproportionation (1.4fold) than those of the wild type

A406V

-

the mutant shows higher thermostability at 50°C, higher intermolecular transglucosylation activity with an upward shift in the optimum temperature and a slight increase in the optimum pH for disproportionation and cyclization reactions compared to the wild type enzyme. The mutant shows higher specific activities for starch transglucosylation (2.8fold) and disproportionation (2.1fold) than those of the wild type

N287Y

-

the mutant shows a significant decrease in all transglucosylation activities including starch transglucosylation, disproportionation, cyclization and coupling compared to the wild type enzyme, while hydrolysis activity is not changed. The mutant shows an increase in thermostability and substrate preference for maltoheptaose in addition to maltotriose

Y172A

-

mutant exhibits lower disproportionation, cyclization, and hydrolysis activities than the wild-type. The kcat/Km of the disproportionation reaction for the Y172A enzyme is 2.8fold lower than that of wild-type. The Y172A enzyme shows a product pattern different from that of wild-type at a long incubation time. The principal large-ring cyclodextrin products of the Y172A mutant are a cycloamylose mixture with a degree of polymerization of 28 or 29

Y418A

-

the mutant shows a significant decrease in starch transglucosylation, disproportionation and cyclization activities compared to the wild type enzyme. The mutant produces large ring-cyclodextrins 36-40 from pea starch

Y418D

-

the mutant shows a significant decrease in starch transglucosylation, disproportionation and cyclization activities compared to the wild type enzyme. The mutant produces large ring-cyclodextrins 36-40 from pea starch

Y418F

-

the mutant shows a significant decrease in starch transglucosylation, disproportionation and cyclization activities compared to the wild type enzyme. The mutant produces large ring-cyclodextrins 29-33 from pea starch

Y418R

-

the mutant shows a significant decrease in starch transglucosylation, disproportionation and cyclization activities compared to the wild type enzyme. The mutant produces large ring-cyclodextrins 36-40 from pea starch

Y418S

-

the mutant shows a significant decrease in starch transglucosylation, disproportionation and cyclization activities compared to the wild type enzyme. The mutant produces large ring-cyclodextrins 36-40 from pea starch

Y418W

-

the mutant shows a significant decrease in starch transglucosylation, disproportionation and cyclization activities compared to the wild type enzyme. The mutant produces large ring-cyclodextrins 36-40 from pea starch

5.5 - 9

-

stable at 30°C

718654

35 - 40

-

at short incubation time for 15 min at 40°C, the remaining activity of the wild type enzyme is 39.5%, whereas for 30 min incubation, the activity remained is 15.2%. At 35°C for a longer incubation time of 3 h, the remaining activity of the wild type enzyme is 45%

35 - 45

-

the specific activity remaining after 10 min at 35°C for the wild type enzyme is 49.3% while that for mutant enzyme N287Y is 91.4%. At 40°C, no activity of the wild type enzyme is left after 10 min but mutant N287Y shows 34.3% remaining activity. At 45°C, the wild type loses all activity after 10 min incubation time while 28.6% of the activity of mutant N287Y still remains

50

-

10 min, almost complete loss of activity

DEAE column chromatography and phenyl column chromatography

HisTrap affinity column chromatography

HisTrap column chromatography

Ni-NTA agarose column chromatography and Superdex S200 gel filtration

HisTrap affinity column chromatography

-

HisTrap affinity column chromatography, and gel filtration

-

HisTrap column chromatography

-

HisTrap column chromatography, and gel filtration

-

expressed in Escherichia coli BL21(DE3) cells

3 entries

expressed in Escherichia coli BL21(DE3)-T1 cells

expressed in Escherichia coli

-

expressed in Escherichia coli BL21(DE3) cells

-

736255, 736324, 755955

expression in Escherichia coli

-

Srisimarat, W.; Kaulpiboon, J.; Krusong, K.; Zimmermann, W.; Pongsawasdi, P.

Altered large-ring cyclodextrin product profile due to a mutation at Tyr-172 in the amylomaltase of Corynebacterium glutamicum

Appl. Environ. Microbiol.

78

7223-7228

2012

Corynebacterium glutamicum

22865069

Srisimarat, W.; Murakami, S.; Pongsawasdi, P.; Krusong, K.

Crystallization and preliminary X-ray crystallographic analysis of the amylomaltase from Corynebacterium glutamicum

Acta Crystallogr. Sect. F

69

1004-1006

2013

Corynebacterium glutamicum (Q8NNA7), Corynebacterium glutamicum

23989149

Rachadech, W.; Nimpiboon, P.; Naumthong, W.; Nakapong, S.; Krusong, K.; Pongsawasdi, P.

Identification of essential tryptophan in amylomaltase from Corynebacterium glutamicum

Int. J. Biol. Macromol.

76

230-235

2015

Corynebacterium glutamicum (Q8NNA7), Corynebacterium glutamicum

25748841

Nimpiboon, P.; Kaulpiboon, J.; Krusong, K.; Nakamura, S.; Kidokoro, S.; Pongsawasdi, P.

Mutagenesis for improvement of activity and thermostability of amylomaltase from Corynebacterium glutamicum

Int. J. Biol. Macromol.

86

820-828

2016

Corynebacterium glutamicum, Corynebacterium glutamicum ATCC 13032

26875536

Vongpichayapaiboon, T.; Pongsawasdi, P.; Krusong, K.

Optimization of large-ring cyclodextrin production from starch by amylomaltase from Corynebacterium glutamicum and effect of organic solvent on product size

J. Appl. Microbiol.

120

912-920

2016

Corynebacterium glutamicum, Corynebacterium glutamicum ATCC 13032

26849173

Tumhom, S.; Krusong, K.; Kidokoro, S.I.; Katoh, E.; Pongsawasdi, P.

Significance of H461 at subsite +1 in substrate binding and transglucosylation activity of amylomaltase from Corynebacterium glutamicum

Arch. Biochem. Biophys.

652

3-8

2018

Corynebacterium glutamicum (Q8NNA7), Corynebacterium glutamicum, Corynebacterium glutamicum DSM 20300 (Q8NNA7)

29885290

Nimpiboon, P.; Krusong, K.; Kaulpiboon, J.; Kidokoro, S.; Pongsawasdi, P.

Roles of N287 in catalysis and product formation of amylomaltase from Corynebacterium glutamicum

Biochem. Biophys. Res. Commun.

478

759-764

2016

Corynebacterium glutamicum, Corynebacterium glutamicum ATCC 13032

27507216

Tumhom, S.; Krusong, K.; Pongsawasdi, P.

Y418 in 410s loop is required for high transglucosylation activity and large-ring cyclodextrin production of amylomaltase from Corynebacterium glutamicum

Biochem. Biophys. Res. Commun.

488

516-521

2017

Corynebacterium glutamicum

28522291

Joo, S.; Kim, S.; Seo, H.; Kim, K.J.

Crystal structure of amylomaltase from Corynebacterium glutamicum

J. Agric. Food Chem.

64

5662-5670

2016

Corynebacterium glutamicum (Q8NNA7), Corynebacterium glutamicum, Corynebacterium glutamicum ATCC 13032 (Q8NNA7)

27366969