CLONING AND MOLECULAR ANALYSIS OF A CDNA AND THE CS-MNP1 GENE ENCODING A MANGANESE PEROXIDASE ISOENZYME FROM THE LIGNIN-DEGRADING BASIDIOMYCETE CERIPORIOPSIS-SUBVERMISPORA

A cDNA (MnP13-1) and the Cs-mnp1 gene encoding for an isoenzyme of man ganese peroxidase (MnP) from C. subvermispora were isolated separately and sequenced. The cDNA, identified in a library constructed in the v ector Lambda ZIPLOX, contains 1285 nucleotides, excluding the poly(A) tail, and has a 63% G + C content. The deduced protein sequence shows a high degree of identity with MnPs from other fungi. The mature prote in contains 364 amino acids, which are preceded by a 24-amino-acid lea der sequence. Consistent with the peroxidase mechanism of MnP, the pro ximal histidine, the distal histidine and the distal arginine are cons erved, although the aromatic binding site (L/V/I-P-X-P) is less hydrop hilic than those of other peroxidases. A gene coding for the same prot ein (Cs-mnp1) was isolated from a genomic library constructed in Lambd a GEM-11 vector using the cDNA MnP13-1 as a probe. A subcloned SacI fr agment of 2.5 kb contained the complete sequence of the Cs-mnp1 gene, including 162 bp and 770 bp of the upstream and downstream regions, re spectively. The Cs-mnp1 gene possesses seven short intervening sequenc es. The intron splice junction sequences as well as the putative inter nal lariat formation sites adhere to the GT-AG and CTRAY rules, respec tively. To examine the structure of the regulatory region of the Cs-mn p1 gene further, a fragment of 1.9 kb was amplified using inverse PCR. A putative TATAA element was identified 5' of the translational start codon. Also, an inverted CCAAT element, SP-1 and AP-2 sites and sever al putative heat-shock and metal response elements were identified. (C ) 1998 Elsevier Science B.V.