Role of dipeptide at extra sugar-binding space of Thermus maltogenic amylase in transglycosylation activity

Jin Sook Baek, Tae J. Kim, Young W. Kim, Hyunju Cha, Jung W. Kim, Yong R. Kim, Sung-Joon Lee, Tae W. Moon, Kwan H. Park

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Two conserved amino acid residues in the extra sugar-binding space near the catalytic site of Thermus maltogenic amylase (ThMA) were analyzed for their role in the hydrolysis and transglycosylation activity of the enzyme. Site-directed mutagenesis was carried out by replacing N331 with a lysine (N331K), E332 with a histidine (E332H), or by replacing both residues at the same time (N331K/E332H). The measured Km values indicated that affinities toward all substrates tested, including starch, pullulan, β-cyclomaltodextrin, and acarbose, were lower in all the mutants compared to that of wild-type ThMA, leading to reduced hydrolysis activity. In addition, the lower ratio of transglycosylation to hydrolysis in the mutants compared to that in the wild-type ThMA indicated that these mutants preferred hydrolysis to the transglycosylation reaction. These results demonstrated that the conserved dipeptide at 331 and 332 of ThMA is directly involved in the formation and accumulation of transfer products by accommodating acceptor sugar molecules.

Original languageEnglish
Pages (from-to)969-975
Number of pages7
JournalJournal of Microbiology and Biotechnology
Volume13
Issue number6
Publication statusPublished - 2003 Dec 1
Externally publishedYes

Fingerprint

glucan 1,4-alpha-maltohydrolase
Thermus
Amylases
Dipeptides
Sugars
Hydrolysis
Acarbose
Mutagenesis
Site-Directed Mutagenesis
Starch
Histidine
Lysine
Amino acids
Catalytic Domain
Enzymes
Amino Acids
Molecules
Substrates

Keywords

  • Acarbose
  • Site-directed mutagenesis
  • Thermus maltogenic amylase (ThMA)
  • Transglycosylation

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Microbiology

Cite this

Baek, J. S., Kim, T. J., Kim, Y. W., Cha, H., Kim, J. W., Kim, Y. R., ... Park, K. H. (2003). Role of dipeptide at extra sugar-binding space of Thermus maltogenic amylase in transglycosylation activity. Journal of Microbiology and Biotechnology, 13(6), 969-975.

Role of dipeptide at extra sugar-binding space of Thermus maltogenic amylase in transglycosylation activity. / Baek, Jin Sook; Kim, Tae J.; Kim, Young W.; Cha, Hyunju; Kim, Jung W.; Kim, Yong R.; Lee, Sung-Joon; Moon, Tae W.; Park, Kwan H.

In: Journal of Microbiology and Biotechnology, Vol. 13, No. 6, 01.12.2003, p. 969-975.

Research output: Contribution to journalArticle

Baek, JS, Kim, TJ, Kim, YW, Cha, H, Kim, JW, Kim, YR, Lee, S-J, Moon, TW & Park, KH 2003, 'Role of dipeptide at extra sugar-binding space of Thermus maltogenic amylase in transglycosylation activity', Journal of Microbiology and Biotechnology, vol. 13, no. 6, pp. 969-975.
Baek, Jin Sook ; Kim, Tae J. ; Kim, Young W. ; Cha, Hyunju ; Kim, Jung W. ; Kim, Yong R. ; Lee, Sung-Joon ; Moon, Tae W. ; Park, Kwan H. / Role of dipeptide at extra sugar-binding space of Thermus maltogenic amylase in transglycosylation activity. In: Journal of Microbiology and Biotechnology. 2003 ; Vol. 13, No. 6. pp. 969-975.
@article{23091355c7364ff088f943bc6f374655,
title = "Role of dipeptide at extra sugar-binding space of Thermus maltogenic amylase in transglycosylation activity",
abstract = "Two conserved amino acid residues in the extra sugar-binding space near the catalytic site of Thermus maltogenic amylase (ThMA) were analyzed for their role in the hydrolysis and transglycosylation activity of the enzyme. Site-directed mutagenesis was carried out by replacing N331 with a lysine (N331K), E332 with a histidine (E332H), or by replacing both residues at the same time (N331K/E332H). The measured Km values indicated that affinities toward all substrates tested, including starch, pullulan, β-cyclomaltodextrin, and acarbose, were lower in all the mutants compared to that of wild-type ThMA, leading to reduced hydrolysis activity. In addition, the lower ratio of transglycosylation to hydrolysis in the mutants compared to that in the wild-type ThMA indicated that these mutants preferred hydrolysis to the transglycosylation reaction. These results demonstrated that the conserved dipeptide at 331 and 332 of ThMA is directly involved in the formation and accumulation of transfer products by accommodating acceptor sugar molecules.",
keywords = "Acarbose, Site-directed mutagenesis, Thermus maltogenic amylase (ThMA), Transglycosylation",
author = "Baek, {Jin Sook} and Kim, {Tae J.} and Kim, {Young W.} and Hyunju Cha and Kim, {Jung W.} and Kim, {Yong R.} and Sung-Joon Lee and Moon, {Tae W.} and Park, {Kwan H.}",
year = "2003",
month = "12",
day = "1",
language = "English",
volume = "13",
pages = "969--975",
journal = "Journal of Microbiology and Biotechnology",
issn = "1017-7825",
publisher = "Korean Society for Microbiolog and Biotechnology",
number = "6",

}

TY - JOUR

T1 - Role of dipeptide at extra sugar-binding space of Thermus maltogenic amylase in transglycosylation activity

AU - Baek, Jin Sook

AU - Kim, Tae J.

AU - Kim, Young W.

AU - Cha, Hyunju

AU - Kim, Jung W.

AU - Kim, Yong R.

AU - Lee, Sung-Joon

AU - Moon, Tae W.

AU - Park, Kwan H.

PY - 2003/12/1

Y1 - 2003/12/1

N2 - Two conserved amino acid residues in the extra sugar-binding space near the catalytic site of Thermus maltogenic amylase (ThMA) were analyzed for their role in the hydrolysis and transglycosylation activity of the enzyme. Site-directed mutagenesis was carried out by replacing N331 with a lysine (N331K), E332 with a histidine (E332H), or by replacing both residues at the same time (N331K/E332H). The measured Km values indicated that affinities toward all substrates tested, including starch, pullulan, β-cyclomaltodextrin, and acarbose, were lower in all the mutants compared to that of wild-type ThMA, leading to reduced hydrolysis activity. In addition, the lower ratio of transglycosylation to hydrolysis in the mutants compared to that in the wild-type ThMA indicated that these mutants preferred hydrolysis to the transglycosylation reaction. These results demonstrated that the conserved dipeptide at 331 and 332 of ThMA is directly involved in the formation and accumulation of transfer products by accommodating acceptor sugar molecules.

AB - Two conserved amino acid residues in the extra sugar-binding space near the catalytic site of Thermus maltogenic amylase (ThMA) were analyzed for their role in the hydrolysis and transglycosylation activity of the enzyme. Site-directed mutagenesis was carried out by replacing N331 with a lysine (N331K), E332 with a histidine (E332H), or by replacing both residues at the same time (N331K/E332H). The measured Km values indicated that affinities toward all substrates tested, including starch, pullulan, β-cyclomaltodextrin, and acarbose, were lower in all the mutants compared to that of wild-type ThMA, leading to reduced hydrolysis activity. In addition, the lower ratio of transglycosylation to hydrolysis in the mutants compared to that in the wild-type ThMA indicated that these mutants preferred hydrolysis to the transglycosylation reaction. These results demonstrated that the conserved dipeptide at 331 and 332 of ThMA is directly involved in the formation and accumulation of transfer products by accommodating acceptor sugar molecules.

KW - Acarbose

KW - Site-directed mutagenesis

KW - Thermus maltogenic amylase (ThMA)

KW - Transglycosylation

UR - http://www.scopus.com/inward/record.url?scp=0346728565&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0346728565&partnerID=8YFLogxK

M3 - Article

AN - SCOPUS:0346728565

VL - 13

SP - 969

EP - 975

JO - Journal of Microbiology and Biotechnology

JF - Journal of Microbiology and Biotechnology

SN - 1017-7825

IS - 6

ER -