Engineering of a manganese-binding site in lignin peroxidase isozyme H8 from Phanerochaete chrysosporium

Tünde Mester, Ming Tien

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

A Mn2+-binding site was created in the recombinant lignin peroxidase isozyme H8 from Phanerochaete chrysosporium. In fungal Mn peroxidase, the Mn-binding site is composed of Glu35, Glu39, and Asp179. We generated a similar site in lignin peroxidase by generating an anionic binding site. We generated three mutations: Asn182Asp, Asp183Lys, and Ala36Glu. Its activity, veratryl alcohol, and Mn2+ oxidation were compared to those of native recombinant enzyme and to fungal Mn peroxidase isozyme H4, respectively. The mutated enzyme was able to oxidize Mn2+ and still retain its ability to oxidize veratryl alcohol. Steadystate results indicate that the enzyme's ability to oxidize veratryl alcohol was lowered slightly. The Km for Mn2+ was determined to be 1.57 mM and the kcat=5.45 s-1. These results indicate that the mutated lignin peroxidase is less effective in Mn2+ oxidation that the wild type fungal enzyme. The pH optima of veratryl alcohol and Mn oxidation were altered by the mutation. They are one unit of pH value higher than those of recombinant H8 and wild type fungal Mn peroxidase isozyme H4.

Original languageEnglish (US)
Pages (from-to)723-728
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume284
Issue number3
DOIs
StatePublished - Jan 1 2001

Fingerprint

manganese peroxidase
Phanerochaete
Manganese
Isoenzymes
Binding Sites
Enzymes
Oxidation
Mutation
veratryl alcohol
lignin peroxidase

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

@article{ed11decedc7548ad9740f856421b8def,
title = "Engineering of a manganese-binding site in lignin peroxidase isozyme H8 from Phanerochaete chrysosporium",
abstract = "A Mn2+-binding site was created in the recombinant lignin peroxidase isozyme H8 from Phanerochaete chrysosporium. In fungal Mn peroxidase, the Mn-binding site is composed of Glu35, Glu39, and Asp179. We generated a similar site in lignin peroxidase by generating an anionic binding site. We generated three mutations: Asn182Asp, Asp183Lys, and Ala36Glu. Its activity, veratryl alcohol, and Mn2+ oxidation were compared to those of native recombinant enzyme and to fungal Mn peroxidase isozyme H4, respectively. The mutated enzyme was able to oxidize Mn2+ and still retain its ability to oxidize veratryl alcohol. Steadystate results indicate that the enzyme's ability to oxidize veratryl alcohol was lowered slightly. The Km for Mn2+ was determined to be 1.57 mM and the kcat=5.45 s-1. These results indicate that the mutated lignin peroxidase is less effective in Mn2+ oxidation that the wild type fungal enzyme. The pH optima of veratryl alcohol and Mn oxidation were altered by the mutation. They are one unit of pH value higher than those of recombinant H8 and wild type fungal Mn peroxidase isozyme H4.",
author = "T{\"u}nde Mester and Ming Tien",
year = "2001",
month = "1",
day = "1",
doi = "10.1006/bbrc.2001.5015",
language = "English (US)",
volume = "284",
pages = "723--728",
journal = "Biochemical and Biophysical Research Communications",
issn = "0006-291X",
publisher = "Academic Press Inc.",
number = "3",

}

Engineering of a manganese-binding site in lignin peroxidase isozyme H8 from Phanerochaete chrysosporium. / Mester, Tünde; Tien, Ming.

In: Biochemical and Biophysical Research Communications, Vol. 284, No. 3, 01.01.2001, p. 723-728.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Engineering of a manganese-binding site in lignin peroxidase isozyme H8 from Phanerochaete chrysosporium

AU - Mester, Tünde

AU - Tien, Ming

PY - 2001/1/1

Y1 - 2001/1/1

N2 - A Mn2+-binding site was created in the recombinant lignin peroxidase isozyme H8 from Phanerochaete chrysosporium. In fungal Mn peroxidase, the Mn-binding site is composed of Glu35, Glu39, and Asp179. We generated a similar site in lignin peroxidase by generating an anionic binding site. We generated three mutations: Asn182Asp, Asp183Lys, and Ala36Glu. Its activity, veratryl alcohol, and Mn2+ oxidation were compared to those of native recombinant enzyme and to fungal Mn peroxidase isozyme H4, respectively. The mutated enzyme was able to oxidize Mn2+ and still retain its ability to oxidize veratryl alcohol. Steadystate results indicate that the enzyme's ability to oxidize veratryl alcohol was lowered slightly. The Km for Mn2+ was determined to be 1.57 mM and the kcat=5.45 s-1. These results indicate that the mutated lignin peroxidase is less effective in Mn2+ oxidation that the wild type fungal enzyme. The pH optima of veratryl alcohol and Mn oxidation were altered by the mutation. They are one unit of pH value higher than those of recombinant H8 and wild type fungal Mn peroxidase isozyme H4.

AB - A Mn2+-binding site was created in the recombinant lignin peroxidase isozyme H8 from Phanerochaete chrysosporium. In fungal Mn peroxidase, the Mn-binding site is composed of Glu35, Glu39, and Asp179. We generated a similar site in lignin peroxidase by generating an anionic binding site. We generated three mutations: Asn182Asp, Asp183Lys, and Ala36Glu. Its activity, veratryl alcohol, and Mn2+ oxidation were compared to those of native recombinant enzyme and to fungal Mn peroxidase isozyme H4, respectively. The mutated enzyme was able to oxidize Mn2+ and still retain its ability to oxidize veratryl alcohol. Steadystate results indicate that the enzyme's ability to oxidize veratryl alcohol was lowered slightly. The Km for Mn2+ was determined to be 1.57 mM and the kcat=5.45 s-1. These results indicate that the mutated lignin peroxidase is less effective in Mn2+ oxidation that the wild type fungal enzyme. The pH optima of veratryl alcohol and Mn oxidation were altered by the mutation. They are one unit of pH value higher than those of recombinant H8 and wild type fungal Mn peroxidase isozyme H4.

UR - http://www.scopus.com/inward/record.url?scp=0034805890&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034805890&partnerID=8YFLogxK

U2 - 10.1006/bbrc.2001.5015

DO - 10.1006/bbrc.2001.5015

M3 - Article

C2 - 11396962

AN - SCOPUS:0034805890

VL - 284

SP - 723

EP - 728

JO - Biochemical and Biophysical Research Communications

JF - Biochemical and Biophysical Research Communications

SN - 0006-291X

IS - 3

ER -