1,4-benzoquinone reductase from Phanerochaete chrysosporium: cDNA cloning and regulation of expression

• Published in: Applied and Environmental Microbiology 1999-02, Vol.65 (2), p.415-421
• Main Authors: Akileswaran, L, Brock, B.J, Cereghino, J.L, Gold, M.H
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; amino acid sequences; Base Sequence; Biological and medical sciences; Biotechnology; Blotting, Northern; Cloning; Cloning, Molecular; complementary DNA; Deoxyribonucleic acid; DNA; DNA, Complementary - analysis; Enzyme Induction; Enzymes; Fundamental and applied biological sciences. Psychology; Fungi; genbank/af106939; gene expression; Gene Expression Regulation, Fungal; Genetic engineering; Genetic technics; Genetics and Molecular Biology; Immunoblotting; messenger RNA; Methods. Procedures. Technologies; Microbiology; Molecular cloning; Molecular Sequence Data; nucleotide sequences; Phanerochaete - enzymology; Phanerochaete - genetics; Phanerochaete chrysosporium; Protein Biosynthesis; Quinone Reductases - chemistry; Quinone Reductases - genetics; Quinone Reductases - metabolism; Sequence Analysis, DNA; Transcription, Genetic
• ISSN: 0099-2240; 1098-5336
• DOI: 10.1128/aem.65.2.415-421.1999

A cDNA clone encoding a quinone reductase (QR) from the white rot basidiomycete Phanerochaete chrysosporium was isolated and sequenced. The cDNA consisted of 1,007 nucleotides and a poly(A) tail and encoded a deduced protein containing 271 amino acids. The experimentally determined eight-amino-acid N-terminal sequence of the purified QR protein from P. chrysosporium matched amino acids 72 to 79 of the predicted translation product of the cDNA. The M(r) of the predicted translation product, beginning with Pro-72, was essentially identical to the experimentally determined M(r) of one monomer of the QR dimer, and this finding suggested that QR is synthesized as a proenzyme. The results of in vitro transcription-translation experiments suggested that QR is synthesized as a proenzyme with a 71-amino-acid leader sequence. This leader sequence contains two potential KEX2 cleavage sites and numerous potential cleavage sites for dipeptidyl aminopeptidase. The QR activity in cultures of P. chrysosporium increased following the addition of 2-dimethoxybenzo-quinone, vanillic acid, or several other aromatic compounds. An immunoblot analysis indicated that induction resulted in an increase in the amount of QR protein, and a Northern blot analysis indicated that this regulation occurs at the level of the qr mRNA.