Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding.

Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding. Research Abstract Details 

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Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding. Abstract Text:

Tatsuya Yamamoto,Shunsuke Izumi,Kunihiko Gekko,

To address the effects of ligand binding on the structural fluctuations of Escherichia coli dihydrofolate reductase (DHFR), the hydrogen/deuterium (H/D) exchange kinetics of its binary and ternary complexes formed with various ligands (folate, dihydrofolate, tetrahydrofolate, NADPH, NADP(+), and methotrexate) were examined using electrospray ionization mass spectrometry. The kinetic parameters of H/D exchange reactions, which consisted of two phases with fast and slow rates, were sensitively influenced by ligand binding, indicating that changes in the structural fluctuation of the DHFR molecule are associated with the alternating binding and release of the cofactor and substrate. No additivity was observed in the kinetic parameters between a ternary complex and its constitutive binary complexes, indicating that ligand binding cooperatively affects the structural fluctuation of the DHFR molecule via long-range interactions. The local H/D exchange profile of pepsin digestion fragments was determined by matrix-assisted laser desorption/ionization mass spectrometry, and the helix and loop regions that appear to participate in substrate binding, largely fluctuating in the apo-form, are dominantly influenced by ligand binding. These results demonstrate that the structural fluctuation of kinetic intermediates plays an important role in enzyme function, and that mass spectrometry on H/D exchange coupled with ligand binding and protease digestion provide new insight into the structure-fluctuation-function relationship of enzymes.

Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding. Publishing Authors By Initials

T Yamamoto,S Izumi,K Gekko,

For similar enzymes and coenzymes: enzymes: oxidoreductases: oxidoreductases acting on ch-nh group donors: tetrahydrofolate dehydrogenase research abstracts see: enzymes and coenzymes: enzymes: oxidoreductases: oxidoreductases acting on ch-nh group donors: tetrahydrofolate dehydrogenase research

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Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding. Journal Published:

PUBLICATION TYPE: Research Support, Non-U.S. Gov

Journal: Journal of biochemistry

VOLUME: 135

Page Numbers: 663-71

Journal Abbreviation: J. Biochem.

ISSN: 0021-924X

DAY: 19

MONTH: Jun

YEAR: 2004

Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding. Information

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LANGUAGE: eng

NlmUniqueID: 376600

Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding. Keywords Mesh Terms:

KEYWORDS: Tetrahydrofolate Dehydrogenase

MESH TERMS: chemistry

Chemical & Substance for Abstract: Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding. Information

Substance Name: Tetrahydrofolate Dehydrogenase

Registry Number: EC 1.5.1.3

Grant and Affiliation Information for Mass spectrometry on hydrogen/deuterium exchange of dihydrofolate reductase: effects of ligand binding.

AFFILIATION: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Higashi-Hiroshima 739-8526, Japan.

Country: Japan

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MEDLINETA: J Biochem

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