A crystallographic and spectroscopic study on the effect of X-ray radiation on the crystal structure of Melanocarpus albomyces laccase

Nina Hakulinen (Corresponding Author), Kristiina Kruus, Anu Koivula, Juha Rouvinen (Corresponding Author)

Research output: Contribution to journalArticleScientificpeer-review

43 Citations (Scopus)

Abstract

Laccases (p-diphenol dioxygen oxidoreductases) belong to the family of blue multicopper oxidases, which catalyse the four-electron reduction of dioxygen to water concomitantly through the oxidation of substrate molecules. Blue multicopper oxidases have four coppers, a copper (T1) forming a mononuclear site and a cluster of three coppers (T2, T3, and T3′) forming a trinuclear site. Because X-rays are known to liberate electrons during data collection and may thus affect the oxidation state of metals, we have investigated the effect of X-ray radiation upon the crystal structure of a recombinant laccase from Melanocarpus albomyces through the use of crystallography and crystal absorption spectroscopy. Two data sets with different strategies, a low and a high-dose data set, were collected at synchrotron. We have observed earlier that the trinuclear site had an elongated electron density amidst coppers, suggesting dioxygen binding. The low-dose synchrotron structure showed similar elongated electron density, but the high-dose X-ray radiation removed the bulk of this density. Therefore, X-ray radiation could alter the active site of laccase from M. albomyces. Absorption spectra of the crystals (320, 420, and 590 nm) during X-ray radiation were measured at a home laboratory. Spectra clearly showed how that the band at 590 nm had vanished, resulting from the T1 copper being reduced, during the long X-ray measurements. The crystal colour changed from blue to colourless. Absorptions at 320 and 420 nm seemed to be rather permanent. The absorption at 320 nm is due to the T3 coppers and it is proposed that absorption at 420 nm is due to the T2 copper when dioxygen or a reaction intermediate is close to this copper.

Original languageEnglish
Pages (from-to)929-934
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume350
Issue number4
DOIs
Publication statusPublished - 2006
MoE publication typeA1 Journal article-refereed

Fingerprint

Laccase
Copper
Crystal structure
X-Rays
Radiation
X rays
Oxygen
Oxidoreductases
Electrons
Synchrotrons
Crystals
Carrier concentration
Reaction intermediates
Oxidation
Crystallography
Absorption spectroscopy
Dosimetry
Absorption spectra
Metals
Color

Keywords

  • Laccase
  • Multicopper oxidase
  • X-ray induced changes
  • Crystal absorption spectroscopy

Cite this

@article{e413773c4f0845f68e5d59c97778d2e3,
title = "A crystallographic and spectroscopic study on the effect of X-ray radiation on the crystal structure of Melanocarpus albomyces laccase",
abstract = "Laccases (p-diphenol dioxygen oxidoreductases) belong to the family of blue multicopper oxidases, which catalyse the four-electron reduction of dioxygen to water concomitantly through the oxidation of substrate molecules. Blue multicopper oxidases have four coppers, a copper (T1) forming a mononuclear site and a cluster of three coppers (T2, T3, and T3′) forming a trinuclear site. Because X-rays are known to liberate electrons during data collection and may thus affect the oxidation state of metals, we have investigated the effect of X-ray radiation upon the crystal structure of a recombinant laccase from Melanocarpus albomyces through the use of crystallography and crystal absorption spectroscopy. Two data sets with different strategies, a low and a high-dose data set, were collected at synchrotron. We have observed earlier that the trinuclear site had an elongated electron density amidst coppers, suggesting dioxygen binding. The low-dose synchrotron structure showed similar elongated electron density, but the high-dose X-ray radiation removed the bulk of this density. Therefore, X-ray radiation could alter the active site of laccase from M. albomyces. Absorption spectra of the crystals (320, 420, and 590 nm) during X-ray radiation were measured at a home laboratory. Spectra clearly showed how that the band at 590 nm had vanished, resulting from the T1 copper being reduced, during the long X-ray measurements. The crystal colour changed from blue to colourless. Absorptions at 320 and 420 nm seemed to be rather permanent. The absorption at 320 nm is due to the T3 coppers and it is proposed that absorption at 420 nm is due to the T2 copper when dioxygen or a reaction intermediate is close to this copper.",
keywords = "Laccase, Multicopper oxidase, X-ray induced changes, Crystal absorption spectroscopy",
author = "Nina Hakulinen and Kristiina Kruus and Anu Koivula and Juha Rouvinen",
year = "2006",
doi = "10.1016/j.bbrc.2006.09.144",
language = "English",
volume = "350",
pages = "929--934",
journal = "Biochemical and Biophysical Research Communications",
issn = "0006-291X",
publisher = "Academic Press",
number = "4",

}

A crystallographic and spectroscopic study on the effect of X-ray radiation on the crystal structure of Melanocarpus albomyces laccase. / Hakulinen, Nina (Corresponding Author); Kruus, Kristiina; Koivula, Anu; Rouvinen, Juha (Corresponding Author).

In: Biochemical and Biophysical Research Communications, Vol. 350, No. 4, 2006, p. 929-934.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - A crystallographic and spectroscopic study on the effect of X-ray radiation on the crystal structure of Melanocarpus albomyces laccase

AU - Hakulinen, Nina

AU - Kruus, Kristiina

AU - Koivula, Anu

AU - Rouvinen, Juha

PY - 2006

Y1 - 2006

N2 - Laccases (p-diphenol dioxygen oxidoreductases) belong to the family of blue multicopper oxidases, which catalyse the four-electron reduction of dioxygen to water concomitantly through the oxidation of substrate molecules. Blue multicopper oxidases have four coppers, a copper (T1) forming a mononuclear site and a cluster of three coppers (T2, T3, and T3′) forming a trinuclear site. Because X-rays are known to liberate electrons during data collection and may thus affect the oxidation state of metals, we have investigated the effect of X-ray radiation upon the crystal structure of a recombinant laccase from Melanocarpus albomyces through the use of crystallography and crystal absorption spectroscopy. Two data sets with different strategies, a low and a high-dose data set, were collected at synchrotron. We have observed earlier that the trinuclear site had an elongated electron density amidst coppers, suggesting dioxygen binding. The low-dose synchrotron structure showed similar elongated electron density, but the high-dose X-ray radiation removed the bulk of this density. Therefore, X-ray radiation could alter the active site of laccase from M. albomyces. Absorption spectra of the crystals (320, 420, and 590 nm) during X-ray radiation were measured at a home laboratory. Spectra clearly showed how that the band at 590 nm had vanished, resulting from the T1 copper being reduced, during the long X-ray measurements. The crystal colour changed from blue to colourless. Absorptions at 320 and 420 nm seemed to be rather permanent. The absorption at 320 nm is due to the T3 coppers and it is proposed that absorption at 420 nm is due to the T2 copper when dioxygen or a reaction intermediate is close to this copper.

AB - Laccases (p-diphenol dioxygen oxidoreductases) belong to the family of blue multicopper oxidases, which catalyse the four-electron reduction of dioxygen to water concomitantly through the oxidation of substrate molecules. Blue multicopper oxidases have four coppers, a copper (T1) forming a mononuclear site and a cluster of three coppers (T2, T3, and T3′) forming a trinuclear site. Because X-rays are known to liberate electrons during data collection and may thus affect the oxidation state of metals, we have investigated the effect of X-ray radiation upon the crystal structure of a recombinant laccase from Melanocarpus albomyces through the use of crystallography and crystal absorption spectroscopy. Two data sets with different strategies, a low and a high-dose data set, were collected at synchrotron. We have observed earlier that the trinuclear site had an elongated electron density amidst coppers, suggesting dioxygen binding. The low-dose synchrotron structure showed similar elongated electron density, but the high-dose X-ray radiation removed the bulk of this density. Therefore, X-ray radiation could alter the active site of laccase from M. albomyces. Absorption spectra of the crystals (320, 420, and 590 nm) during X-ray radiation were measured at a home laboratory. Spectra clearly showed how that the band at 590 nm had vanished, resulting from the T1 copper being reduced, during the long X-ray measurements. The crystal colour changed from blue to colourless. Absorptions at 320 and 420 nm seemed to be rather permanent. The absorption at 320 nm is due to the T3 coppers and it is proposed that absorption at 420 nm is due to the T2 copper when dioxygen or a reaction intermediate is close to this copper.

KW - Laccase

KW - Multicopper oxidase

KW - X-ray induced changes

KW - Crystal absorption spectroscopy

U2 - 10.1016/j.bbrc.2006.09.144

DO - 10.1016/j.bbrc.2006.09.144

M3 - Article

VL - 350

SP - 929

EP - 934

JO - Biochemical and Biophysical Research Communications

JF - Biochemical and Biophysical Research Communications

SN - 0006-291X

IS - 4

ER -