Indigo production by naphthalene-degrading bacteria

Abstract

A I N . 2000. A wild-type naphthalene-degrading strain Pseudomonas putida RKJ1 and two recombinant strains each of Ps. putida and Escherichia coli carrying the genes for naphthalene degradation on a recombinant plasmid pRKJ3, produced indigo and indirubin pigments from indole. Naphthalene, salicylate and IPTG induced cells of naphthalene-degrading recombinant bacteria produced up to two times higher indigo compared with the uninduced cells. The maximum rates of indigo formation by Ps. putida RKJ1, Ps. putida RKJ5/pRKJ3, Ps. putida KT2442/pRKJ3, E. coli TB1/pRKJ3 and E. coli AB1157/pRKJ3 were 0Á60, 0Á80, 0Á60, 1Á20 and 1Á50 nmol min À1 mg dry biomass À1 , respectively, using indole as the substrate. The apparent K m values of indigo formation by these same bacteria were 0Á22, 0Á15, 0Á10, 0Á21 and 0Á20 mmol l À1 , respectively, again using indole as the substrate. The present study revealed that E. coli AB1157 was the most ef®cient of the hosts tested for the expression of the plasmid encoded genes (pRKJ3) from the wild-type strain Ps. putida RKJ1. In addition, both recombinant E. coli strains were capable of producing indigo directly from nutrient medium. INTRODUCTIONIndigo is one of the oldest textile dyes and was traditionally produced from an extract of the plants of the genus Indigofera. Presently, it is being produced by chemical synthesis and is marketed by many leading companies. Very few reports are available regarding the microbial biosynthesis of indigo. Among the microbial indigo producers, the majority of micro-organisms are aromatic hydrocarbondegrading bacteria such as naphthalene-degrading Pseudomonas putida NDO (Murdock et al. 1993), Ps. putida PpG7 (O'Connor and Hartmans 1998), p-cumate and mand p-toluate-degrading Ps. putida F1 and Ps. putida mt-2, respectively (Eaton and Chapman 1995), styrene-degrading Ps. putida S12 and CA-3 (O'Connor et al. 1997) and toluene-degrading Ps. mendocina KR1 (Yen et al. 1991). The enzyme system responsible for indigo formation generally consists of one or more enzymes, typically monooxygenases, dioxygenases or hydroxylases (O'Connor et al. 1997;Doukyu et al. 1998). The genes encoding these enzymes have been cloned from Pseudomonas and Rhodococcus spp. into Escherichia coli strains (Yen et al. 1991;Hart et al. 1992;Eaton and Chapman 1995) so as to produce indigo directly from either nutrient medium (Ensley et al. 1983) or from glucose (Murdock et al. 1993).Recently, a method has been patented by O'riel and Kim (1998) for producing indigo and indirubin dyes using a recombinant Escherichia coli containing a gene encoding a phenol hydroxylase from Bacillus stearothermophilus. In the present study, we report on the formation and identi®ca-tion of indigo and indirubin pigments by one wild-type and four recombinant bacterial strains. MATERIALS AND METHODS Bacteria, media and culture conditionsOf the ®ve bacteria used in the present study, Ps. putida RKJ1 (Samanta et al. 1998) was a wild-type strain and was capable of degrading naphthalene through salicylate. Four recombi...

Indigo biosynthesis by Comamonas sp. MQ

“…peaks at m/z 148 and 263 with retention times of 3.5 and 9.9 min were observed (Supplementary information), which exactly matched the fragmentation pattern of commercial isatin and indigo standards, respectively. Compared with fragmentation pattern and UV-visible spectra in previous reports, it further confirmed that these two compounds were isatin (3.5 min) and indigo (9.9 min) (O'Connor et al 1997;Bhushan et al 2000;Meyer et al 2002).…”

“…and Acinetobacter spp. (O'Connor and Hartmans 1998;Bhushan et al 2000;Qu et al 2010). However, indigo biosynthesis from indole by Comamonas sp.…”

Hydroxylation of Indole by Laboratory-evolved 2-Hydroxybiphenyl 3-Monooxygenase

“…Naphthalene dioxygenase was soon found not to be the only enzyme capable of indole oxidation: several other oxygenases that accept indole as a substrate have been identified (5)(6)(7)(8)(9)(10). These are either enzymes similar to naphthalene dioxygenase that activate oxygen with iron centers or members of the cytochrome P450 family (6,11).…”

Biosynthesis of Indigo Dye by Newly Isolated Naphthalene-Degrading Strain Pseudomonas sp. HOB1 and its Application in Dyeing Cotton Fabric

“…HOB1 was found to be 77.8 nmol min −1 mg dry biomass −1 . This rate of indigo formation is 50-100 times higher as compared to the indigo formation rates (0.60, 0.80, 1.2, and 1.5 nmol min −1 mg dry biomass −1 ) reported by Bhushan et al [19] under different inducing conditions with wild-type Pseudomonas putida strain as well as [16,20] have reported indigo formation rate of 8.3 and 12 nmol min −1 mg dry biomass −1 , respectively, which is again much less than indigo formation rate by Pseudomonas sp. HOB1.…”