Cloning and expression of thermophilic catechol 1,2-dioxygenase gene (catA) fromStreptomyces setonii

• DOI: 10.1111/j.1574-6968.2001.tb10491.x
• Publication Date: 2006-01-09T14:48:12Z
• Authors: Hae-Reun An; Hyun-Joo Park; Eung-Soo Kim
• Keywords: Heterologous expression

Streptomyces setonii (ATCC 39116) degrades various single aromatic compounds such as phenol or benzoate via an ortho-cleavage pathway using catechol 1,2-dioxygenase (C12O). A PCR degenerate primers based on the conserved regions of known C12O-encoding genes amplified a 0.45-kbp DNA fragment from S. total DNA. Southern hybridization analysis and size-selected library screening product probe led to isolation 6.4-kbp fragment, which genetic locus was found be located within 1.4-kbp fragment. complete nucleotide sequencing revealed 0.84-kbp open reading frame, showed strong overall amino acid similarity high-G+C Gram-positive (but significantly less Gram-negative) bacterial mesophilic C12Os. The heterologous expression cloned in Escherichia coli demonstrated that this C12O possessed thermophilic activity broad temperature range (up 65°C) higher against 3-methylcatechol than 4-methylcatechol, but no protocatechuate.