Transcript and activity levels of different Pleurotus ostreatus peroxidases are differentially affected by Mn2+.
The white-rot fungus Pleurotus ostreatus produces both manganese-dependent peroxidase (MnP) and versatile peroxidase (VP) in non-manganese-amended peptone medium (PM). We studied the effect of Mn2+ supplementation on MnPs and VPs in P. ostreatus by analysing the enzymatic and transcript abundance profiles of the peroxidases, as well as the lignin mineralization rate. The fungus was grown in PM under solid-state conditions using perlite as an inert solid support. Mn2+ amendment resulted in a 1.7-fold increase in [14C]-lignin mineralization relative to unamended medium. Anion-exchange chromatography was used to resolve the fungal peroxidase's enzymatic activity profile. Five peaks (P1-P5) of VP and one peak (P6) of MnP activity were detected in unamended medium. In Mn2+-amended medium, a reduction in the activity of the VPs was observed. On the other hand, a sharp increase in the MnP activity level of peak P6 was detected. The P6 isoenzyme was purified and showed manganese-dependent peroxidation of phenolic substrates. Internal sequence analysis of the purified enzyme revealed 100% identity with the deduced amino acid sequence of P. ostreatus MnP3 (GenBank AB016519). The effect of Mn2+ on the relative abundance of gene transcripts of three VPs and one MnP from P. ostreatus was monitored using reverse transcription-polymerase chain reaction (RT-PCR) with oligonucleotide primer sets synthesized on the basis of non-conserved sequences of the different peroxidases. The reduction in VP gene transcript abundance and the increase in mnp3 transcript level were collinear with the changes observed in the enzyme activity profiles. These results indicate that the activity of peroxidases is regulated at the transcriptional level. We suggest that the expression of MnP and VP may be differentially regulated by the presence of Mn2+.
Cohen
R
R
Department of Plant Pathology and Microbiology and The Otto Warburg Center for Biotechnology in Agriculture, Faculty of Agricultural, Food and Environmental Quality Sciences, The Hebrew University of Jerusalem, PO Box 12, Rehovot 76100, Israel.
Hadar
Y
Y
Yarden
O
O
D000363:Agaricales / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D003470:Culture Media; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects*; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008031:Lignin / Q000378:metabolism; D008345:Manganese / Q000494:pharmacology*; D008969:Molecular Sequence Data; D010544:Peroxidases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D014158:Transcription, Genetic / Q000187:drug effects*
D003470:Culture Media; D007527:Isoenzymes; D012333:RNA, Messenger; D008345:Manganese; D008031:Lignin; D010544:Peroxidases