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

Abstract

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

Four Aromatic Intradiol Ring Cleavage Dioxygenases from Aspergillus niger

“…For example, as shown in Fig. 5a, spectral changes following cleavage of catechol by NRRL3_02644 leads to the formation of cis-cis muconic acid, as indicated by the increase of absorbance between 220 and 300 nm, with the highest absorbance observed at 260 nm (22). Similar spectral changes in the UV region of the spectrum were observed after the addition of NRRL3_02644 to the methyl-substituted catechols 4-methylcatechol and 3-methylcatechol.…”

Draft Genome Sequence of Streptomyces sp. Strain NL15-2K, a Degrader of Lignin-Derived Aromatic Compounds, Isolated from Forest Soil

“…The antiSMASH algorithm predicted gene clusters for the biosynthesis of coelichelin, alkylresorcinol, gamma-butyrolactone, albaflavenone, desferrioxamine B, ectoine, endophenazines, indigoidine, and GE37468. Moreover, the genome contained genes involved in the catabolism of lignin-derived aromatic compounds, including pcaHG (14) and catA (15), which encode protocatechuate 3,4-dioxygenase and catechol 1,2-dioxygenase, respectively, and play a role in cleavage of the aromatic ring. Thus, this study provides valuable genetic information required to understand the catabolism of lignin-derived aromatic compounds in strain NL15-2K and to develop biocatalysts for producing valuable compounds from inexpensive plant constituents.…”

Identification and characterization of cresol degrading Pseudomonas monteilii strain SHY from Soil samples

“…Initially, hydroxylase act on the phenolic ring forming catechol followed by oxidation resulting into ring opening [13], [14]. The oxidation is basically of two types ortho and meta, where, two different types of enzymes are involved, catechol 1,3 dioxygenase and catechol 2,3 dioxygenase, acting respectively [10], [15]. The ortho-oxidation pathway in degradation process is also referred to as beta-ketoadipate pathway [2],[16].…”