Anthranilate degradation by an eurypsychrophilic Pseudomonas sp. form Alps

Abstract

The new alpine soil bacterium Pseudomonas sp. strain KOPRI 25931 was selected for the excellent ability to grow on anthranilate (2-aminobenzoate) and benzoate from Polar and Alpine Microbial Collection (PAMC) in Korea Polar Research Institute (KOPRI). It was characterized to be eurypsychrophilic (both psychrophilic and mesotolerant) with a broad temperature range of 5  30oC for benzoate degradation and cell growth. Except anthranilate and benzoate, KOPRI 25931 was capable of growth on only catechol among mono- substituted benzoates and central metabolites tested. Crude lysate from anthranilate- grown KOPRI 25931 cells showed catechol 1,2-dioxygenase activity against catechol. The genes encoding the initial steps in anthranilate metabolism were cloned in a fosmid clone, sequenced, and analyzed to reveal the six catabolic and two regulatory genes (catACBR and antRABC), which are tightly clustered within an approximately 7.8 kb-region on chromosome. When expressed in Escherichia coli, AntABC (anthranilate 1,2- dioxygenase) transformed anthranilate into catechol, and CatA (catechol 1,2-dioxygenase) showed higher ortho-cleavage activity against catechol than others. The growth test, metabolite identification, and enzyme assay demonstrate that AntABC in KOPRI 25931 oxidizes anthranilate to catechol, which is further degraded to cis,cis-muconate through ortho-cleavage pathway initi