The crystal structure of lignin peroxidase at 1.70 Å resolution reveals a hydroxy group on the Cβ of tryptophan 171:: A novel radical site formed during the redox cycle

被引:139
作者
Choinowski, T [1 ]
Blodig, W [1 ]
Winterhalter, KH [1 ]
Piontek, K [1 ]
机构
[1] Swiss Fed Inst Technol, Biochem Lab 1, CH-8092 Zurich, Switzerland
关键词
lignin peroxidase; crystal structure; redox cycle; protein radical; redox potential;
D O I
10.1006/jmbi.1998.2507
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The crystal structure of lignin peroxidase (LiP) from the white rot fungus Phanerochaete chrysosporium was refined to an X-factor of 16.2% utilizing synchrotron data in the resolution range from 10 to 1.7 Angstrom. The final model comprises all 343 amino acid residues, 370 water molecules, the heme, four carbohydrates, and two calcium ions. Lignin peroxidase shows the typical peroxidase fold and the heme has a close environment as found in other peroxidases. During refinement of the Lip model an unprecedented modification of an amino acid was recognized. The surface residue tryptophan 171 in LiP is stereospecifically hydroxylated at the CP atom due to an autocatalytic process. We propose that during the catalytic cycle of Lip a transient radical at Trp171 occurs that is different from those previously assumed for this type of peroxidase. Recently, the existence of a second substrate-binding site centered at Trp171 has been reported, by us which is different from the "classical heme edge" site found in other peroxidases. Here, we report evidence for a radical formation at Trp171 using spin trapping, which supports the concept of Trp171 being a redox active amino acid and being involved in the oxidation of veratryl alcohol. On the basis of our current model, an electron pathway from Trp171 to the heme is envisaged, relevant for the oxidation of veratryl alcohol and possibly lignin. Beside the opening leading to the heme edge, which can accommodate small aromatic substrate molecules, a smaller channel giving access to the distal heme pocket was identified that is large enough for molecules such as hydrogen peroxide. Furthermore, it was found that in Lip the bond between the heme iron and the N-epsilon 2 atom of the proximal histidine residue is significantly longer than in cytochrome c peroxidase (CcP). The weaker Fe-N bond in LiP renders the heme more electron deficient and destabilizes high oxidation states, which could explain the higher redox potential of Lip as compared to CcP. (C) 1999 Academic Press.
引用
收藏
页码:809 / 827
页数:19
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共 82 条
[1]  
ABOLA EE, 1987, CRYSTALLOGRAPHIC DAT, P107
[2]   LIGNIN CHEMISTRY - PAST, PRESENT AND FUTURE [J].
ADLER, E .
WOOD SCIENCE AND TECHNOLOGY, 1977, 11 (03) :169-218
[3]   ENZYME-CATALYZED OXIDATION OF NONPHENOLIC AROMATIC-COMPOUNDS [J].
AGEORGES, A ;
PELTER, A ;
WARD, RS .
PHYTOCHEMISTRY, 1991, 30 (01) :121-126
[4]   LIGNIN PEROXIDASE - RESONANCE RAMAN SPECTRAL EVIDENCE FOR COMPOUND-II AND FOR A TEMPERATURE-DEPENDENT COORDINATION-STATE EQUILIBRIUM IN THE FERRIC ENZYME [J].
ANDERSSON, LA ;
RENGANATHAN, V ;
LOEHR, TM ;
GOLD, MH .
BIOCHEMISTRY, 1987, 26 (08) :2258-2263
[5]   THE CCP4 SUITE - PROGRAMS FOR PROTEIN CRYSTALLOGRAPHY [J].
BAILEY, S .
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY, 1994, 50 :760-763
[6]   PROTON NMR INVESTIGATION INTO THE BASIS FOR THE RELATIVELY HIGH REDOX POTENTIAL OF LIGNIN PEROXIDASE [J].
BANCI, L ;
BERTINI, I ;
TURANO, P ;
TIEN, M ;
KIRK, TK .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1991, 88 (16) :6956-6960
[7]   AMINO-ACID-SEQUENCE OF COPRINUS-MACRORHIZUS PEROXIDASE AND CDNA SEQUENCE ENCODING COPRINUS-CINEREUS PEROXIDASE - A NEW FAMILY OF FUNGAL PEROXIDASES [J].
BAUNSGAARD, L ;
DALBOGE, H ;
HOUEN, G ;
RASMUSSEN, EM ;
WELINDER, KG .
EUROPEAN JOURNAL OF BIOCHEMISTRY, 1993, 213 (01) :605-611
[8]   PROTEIN DATA BANK - COMPUTER-BASED ARCHIVAL FILE FOR MACROMOLECULAR STRUCTURES [J].
BERNSTEIN, FC ;
KOETZLE, TF ;
WILLIAMS, GJB ;
MEYER, EF ;
BRICE, MD ;
RODGERS, JR ;
KENNARD, O ;
SHIMANOUCHI, T ;
TASUMI, M .
JOURNAL OF MOLECULAR BIOLOGY, 1977, 112 (03) :535-542
[9]   Autocatalytic formation of a hydroxy group at Cβ of Trp171 in lignin peroxidase [J].
Blodig, W ;
Doyle, WA ;
Smith, AT ;
Winterhalter, K ;
Choinowski, T ;
Piontek, K .
BIOCHEMISTRY, 1998, 37 (25) :8832-8838
[10]   CRYSTALLOGRAPHIC R-FACTOR REFINEMENT BY MOLECULAR-DYNAMICS [J].
BRUNGER, AT ;
KURIYAN, J ;
KARPLUS, M .
SCIENCE, 1987, 235 (4787) :458-460