The Km for FAD was determined to be 47 μM The enzyme catalyzed

The Km for FAD was determined to be 4.7 μM. The enzyme catalyzed the conversion of 1-H2NA to 1,2-DHN only under aerobic conditions. These results suggested that 1-hydroxy-2-naphthoic acid hydroxylase is a flavoprotein monooxygenase specific for 1-H2NA and different from salicylate-1-hydroxylase. In bacteria, phenanthrene is metabolized to a key intermediate, 1-hydroxy-2-naphthoic acid (1-H2NA),

which is further channelized to the central carbon pathway either via a ‘naphthalene route’ (Rogoff & Wender, 1957; Evans et al., 1965; Prabhu & Phale, 2003) or via a ‘phthalate route’ (Iwabuchi & Harayama, 1998; Deveryshetty, 2009; Deveryshetty & Phale, 2009). In the ‘naphthalene route’, 1-H2NA is metabolized via 1,2-dihydroxynaphthalene (1,2-DHN) selleck chemicals and salicylic acid

to catechol by a series of enzymatic steps similar to naphthalene metabolic pathway. Biochemical and genetic studies suggest that enzymes responsible for the conversion of naphthalene to salicylic acid could also transform find more phenanthrene to 1-H2NA (Menn et al., 1993; Kiyohara et al., 1994; Takizawa et al., 1994). Phenanthrene-degrading Pseudomonas putida strain BS202P1, which also metabolizes naphthalene, is reported to have a broad substrate-specific salicylate-1-hydroxylase which is also responsible for the conversion of 1-H2NA to 1,2-DHN (Balashova et al., 2001). However, the enzyme showed a higher catalytic from efficiency for salicylic acid as compared to 1-H2NA. N-terminal amino acid sequence

showed significant homology with salicylate-1-hydroxylases from other gram-negative bacteria (Balashova et al., 2001). Soil isolate Alcaligenes sp. strain PPH degrades phenanthrene as the sole carbon source. The specific activity versus growth profile indicated the presence of two hydroxylases, salicylate-1-hydroxylase and 1-hydroxy-2-naphthoic acid hydroxylase, in this strain (Deveryshetty, 2009). Salicylate-1-hydroxylase from various bacterial sources have been characterized and reported to have wide substrate specificity (Yamamoto et al., 1965; Katagiri et al., 1966; White-Stevens et al., 1972; Tu et al., 1981; You & Roe, 1981; You et al., 1990; Bosch et al., 1999; Balashova et al., 2001; Pinyakong et al., 2003; Zhao et al., 2005; Jouanneau et al., 2007). The hydroxylation of 1-H2NA to 1,2-DHN is similar to that of salicylic acid to catechol. However, the enzyme specific for 1-H2NA has not been reported so far. The aim of the present study is to purify 1-hydroxy-2-naphthoic acid hydroxylase from Alcaligenes sp. strain PPH and study its kinetic properties and substrate specificity. Here, we report partial purification and characterization of 1-hydroxy-2-naphthoic acid hydroxylase from the phenanthrene-degrading Alcaligenes sp. strain PPH.

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