Phenol/cresol degradation by the thermophilic Bacillus thermoglucosidasius A7: cloning and sequence analysis of five genes involved in the pathway.
Bacillus thermoglucosidasius A7 degraded phenol at 65 degrees C via the meta cleavage pathway. Five enzymes used in the metabolism of phenol were cloned from B. thermoglucosidasius A7 into pUC18. Nine open reading frames were present on the 8.1kb insert, six of which could be assigned a function in phenol degradation using database homologies and enzyme activities. The phenol hydroxylase is a two-component enzyme encoded by pheA1 and pheA2. The larger component (50kDa) has 49% amino acid identity with the 4-hydroxyphenylacetate hydroxylase of Escherichia coli, while the smaller component (19kDa) is most related (30% amino acid identity) to the styrene monoxygenase component B from Pseudomonas fluorescens. Both components were neccessary for activity. The catechol 2, 3-dioxygenase encoded by pheB has 45% amino acid identity with dmpB of Pseudomonas sp. CF600 and could be assigned to superfamily I, family 2 and a new subfamily of the Eltis and Bolin grouping. The 2-hydroxymuconic acid semialdehyde hydrolase (2HMSH), encoded by pheC, revealed the highest amino acid identity (36%) to the equivalent enzyme from Pseudomonas sp. strain CF600, encoded by dmpD. Based on sequence identity, pheD and pheE were deduced to encode the 2-hydroxypenta-2,4-dienoate hydratase (2HDH), demonstrating 45% amino acid identity to the gene product of cumE from Pseudomonas fluorescens and the acetaldehyde dehydrogenase (acylating) demonstrating 57% amino acid identity to the gene product of bphJ from Pseudomonas LB400.
Duffner
F M
FM
Biotechnology II, Technical University Hamburg-Harburg, Denickestr. 15, 21071, Hamburg, Germany. fiod@novo.dk
Kirchner
U
U
Bauer
M P
MP
Müller
R
R
D000445:Aldehyde Oxidoreductases / Q000235:genetics; D001407:Bacillus / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D001673:Biodegradation, Environmental; D050557:Catechol 2,3-Dioxygenase; D003001:Cloning, Molecular; D003408:Cresols / Q000378:metabolism*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D049308:Dioxygenases; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics; D006836:Hydro-Lyases / Q000235:genetics; D006867:Hydrolases / Q000235:genetics; D006899:Mixed Function Oxygenases / Q000235:genetics; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D009876:Operon; D010105:Oxygenases / Q000235:genetics; D019800:Phenol / Q000378:metabolism*; D011506:Proteins / Q000235:genetics; D017422:Sequence Analysis, DNA
D003408:Cresols; D004269:DNA, Bacterial; D011506:Proteins; D019800:Phenol; D006899:Mixed Function Oxygenases; D010105:Oxygenases; D049308:Dioxygenases; D050557:Catechol 2,3-Dioxygenase; C022336:phenol 2-monooxygenase; D000445:Aldehyde Oxidoreductases; C018699:aldehyde dehydrogenase (NAD(P)+); D006867:Hydrolases; C092541:2-hydroxymuconate-semialdehyde hydrolase; C098714:2-hydroxypenta-2,4-dienoate hydratase; D006836:Hydro-Lyases; C077977:cresol