Functional assignment of Glu386 and Arg388 in the active site of l-galactono-¿-lactone dehydrogenase

N.G.H. Leferink; M.D.F. Jose; W.A.M. van den Berg; W.J.H. van Berkel

doi:10.1016/j.febslet.2009.09.004

Functional assignment of Glu386 and Arg388 in the active site of l-galactono-¿-lactone dehydrogenase

N.G.H. Leferink, M.D.F. Jose, W.A.M. van den Berg, W.J.H. van Berkel

Research output: Contribution to journal › Article › Academic › peer-review

22 Citations (Scopus)

Abstract

The flavoenzyme l-galactono-¿-lactone dehydrogenase (GALDH) catalyzes the terminal step of vitamin C biosynthesis in plants. Little is known about the catalytic mechanism of GALDH and related aldonolactone oxidoreductases. Here we identified an essential Glu–Arg pair in the active site of GALDH from Arabidopsis thaliana. Glu386 and Arg388 variants show high Km values for l-galactono-1,4-lactone and low turnover rates. Arg388 is crucial for the stabilization of the anionic form of the reduced FAD cofactor. Glu386 is involved in productive substrate binding. The E386D variant has lost its specificity for l-galactono-1,4-lactone and shows the highest catalytic efficiency with l-gulono-1,4-lactone

Original language	English
Pages (from-to)	3199-3203
Journal	FEBS Letters
Volume	583
Issue number	19
DOIs	https://doi.org/10.1016/j.febslet.2009.09.004
Publication status	Published - 2009

Keywords

l-galactono-1,4-lactone dehydrogenase
flavocytochrome b(2)
cholesterol oxidase
oxygen reactivity
ascorbic-acid
vitamin-c
complex i
biosynthesis
plant
arabidopsis

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@article{e86d6f631d2546e3ba19ac5b5c3ed1b0,

title = "Functional assignment of Glu386 and Arg388 in the active site of l-galactono-¿-lactone dehydrogenase",

abstract = "The flavoenzyme l-galactono-¿-lactone dehydrogenase (GALDH) catalyzes the terminal step of vitamin C biosynthesis in plants. Little is known about the catalytic mechanism of GALDH and related aldonolactone oxidoreductases. Here we identified an essential Glu–Arg pair in the active site of GALDH from Arabidopsis thaliana. Glu386 and Arg388 variants show high Km values for l-galactono-1,4-lactone and low turnover rates. Arg388 is crucial for the stabilization of the anionic form of the reduced FAD cofactor. Glu386 is involved in productive substrate binding. The E386D variant has lost its specificity for l-galactono-1,4-lactone and shows the highest catalytic efficiency with l-gulono-1,4-lactone",

keywords = "l-galactono-1,4-lactone dehydrogenase, flavocytochrome b(2), cholesterol oxidase, oxygen reactivity, ascorbic-acid, vitamin-c, complex i, biosynthesis, plant, arabidopsis",

author = "N.G.H. Leferink and M.D.F. Jose and {van den Berg}, W.A.M. and {van Berkel}, W.J.H.",

year = "2009",

doi = "10.1016/j.febslet.2009.09.004",

language = "English",

volume = "583",

pages = "3199--3203",

journal = "FEBS Letters",

issn = "0014-5793",

publisher = "Wiley",

number = "19",

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TY - JOUR

T1 - Functional assignment of Glu386 and Arg388 in the active site of l-galactono-¿-lactone dehydrogenase

AU - Leferink, N.G.H.

AU - Jose, M.D.F.

AU - van den Berg, W.A.M.

AU - van Berkel, W.J.H.

PY - 2009

Y1 - 2009

N2 - The flavoenzyme l-galactono-¿-lactone dehydrogenase (GALDH) catalyzes the terminal step of vitamin C biosynthesis in plants. Little is known about the catalytic mechanism of GALDH and related aldonolactone oxidoreductases. Here we identified an essential Glu–Arg pair in the active site of GALDH from Arabidopsis thaliana. Glu386 and Arg388 variants show high Km values for l-galactono-1,4-lactone and low turnover rates. Arg388 is crucial for the stabilization of the anionic form of the reduced FAD cofactor. Glu386 is involved in productive substrate binding. The E386D variant has lost its specificity for l-galactono-1,4-lactone and shows the highest catalytic efficiency with l-gulono-1,4-lactone

AB - The flavoenzyme l-galactono-¿-lactone dehydrogenase (GALDH) catalyzes the terminal step of vitamin C biosynthesis in plants. Little is known about the catalytic mechanism of GALDH and related aldonolactone oxidoreductases. Here we identified an essential Glu–Arg pair in the active site of GALDH from Arabidopsis thaliana. Glu386 and Arg388 variants show high Km values for l-galactono-1,4-lactone and low turnover rates. Arg388 is crucial for the stabilization of the anionic form of the reduced FAD cofactor. Glu386 is involved in productive substrate binding. The E386D variant has lost its specificity for l-galactono-1,4-lactone and shows the highest catalytic efficiency with l-gulono-1,4-lactone

KW - l-galactono-1,4-lactone dehydrogenase

KW - flavocytochrome b(2)

KW - cholesterol oxidase

KW - oxygen reactivity

KW - ascorbic-acid

KW - vitamin-c

KW - complex i

KW - biosynthesis

KW - plant

KW - arabidopsis

U2 - 10.1016/j.febslet.2009.09.004

DO - 10.1016/j.febslet.2009.09.004

M3 - Article

SN - 0014-5793

VL - 583

SP - 3199

EP - 3203

JO - FEBS Letters

JF - FEBS Letters

IS - 19

ER -

Functional assignment of Glu386 and Arg388 in the active site of l-galactono-¿-lactone dehydrogenase

Abstract

Keywords

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