Microbial degradation kinetics and molecular mechanism of 2,6-dichloro-4-nitrophenol by a Cupriavidus strain

TLDR: CNP-8 is the first bacterium with the ability to utilize 2,6-DCNP, and this study fills a gap in the microbial degradation mechanism of this pollutant at the molecular, biochemical and genetic levels.

About: This article is published in Environmental Pollution.The article was published on 2020-03-01 and is currently open access. It has received 12 citations till now. The article focuses on the topics: Cupriavidus & Microbial biodegradation.

Figures

Fig. 6. Enzymatic activity assay of HnpAB against 2,6-DCNP, together with the products identification. (A) Spectral changes during the transformation of 2,6-CBNP by purified HnpAB. (B) Spectral changes during the transformation of 2,6-CBNP by only purified HnpA. (C) The mass spectra of acetylated 2,6-DCHQ. (D) The mass spectra of acetylated 6-CHQ.

Fig. 7. (A) Phylogenetic relationship of the oxygenase component of the two-component phenols monooxygenases including HnpA. HnpA is indicated by a “+”. (B) Comparison of the genetic organization of the hnp gene cluster involved in 2,6-DCNP catabolism in strain CNP-8, the tcp gene cluster involved in 2,4,6-TCP catabolism in Cuptiavidus necator JMP134 and the gene clusters involved in 4-nitrophenol catabolism in some bacteria.

Fig. 1. Time course of 2,6-DCNP degradation by Cupriavidus sp. strain CNP-8 in mineral salts medium, together with the accumulation of NO2 , Cl and OD600. (▪) 2,6-DCNP concentration with strain CNP-8 inoculation; (,) 2,6-DCNP concentration without strain CNP-8 inoculation; (C) OD600 with 2,6-DCNP addition; (B) OD600 without 2,6- DCNP addition; (:) NO2 (;) Cl .

Fig. 8. The catabolic pathway of 2,6-DCNP in Cupriavidus sp. strain CNP-8. TCA, tricarboxylic acid. The unidentified intermediate is in bracket.

Fig. 2. Experimental and predicted specific growth rates of strain CNP-8 against 2,6- DCNP according to Haldane’s model. Fig. 3. Simultaneous degradation of three chlorinated nitrophenols by strain CNP-8 from the synthetic wastewater. (▪) 2C4NP; (C 2C5NP; (:) 2,6-DCNP.

Fig. 4. Biotransformation of 2,6-DCNP (A) and 2,6-DBNP (B) by the induced and uninduced cells of strain CNP-8. (,) uninduced cells; (B) 2,6-DCNP-induced cells; (△) 2,6-DBNPinduced cells.

Frequently asked questions

Q1. What have the authors contributed in "Microbial degradation kinetics and molecular mechanism of 2,6-dichloro-4-nitrophenol by a cupriavidus strain" ?

However, microorganisms with the ability to utilize 2,6DCNP have not been reported. In this study, Cupriavidus sp. CNP-8 having been previously reported to degrade various halogenated nitrophenols, was verified to be also capable of degrading 2,6-DCNP. To their knowledge, CNP-8 is the first bacterium with the ability to utilize 2,6-DCNP, and this study fills a gap in the microbial degradation mechanism of this pollutant at the molecular, biochemical and genetic levels. Moreover, strain CNP-8 could degrade three chlorinated nitrophenols rapidly from the synthetic wastewater, indicating its potential in the bioremediation of chlorinated nitrophenols polluted environments. 

Q2. How many mM chlorine ion was accumulated when 2,6-DCNP was?

0.76 mM chlorine ion was accumulated when 2,6-DCNP was completely degraded, indicating that strain CNP-8 can remove two chlorines from 2,6-DCNP. 

Q3. How many cells per liter was the strain CNP-8 cultivated overnight?

Strain CNP-8was initially cultivated overnight in LB medium and washed twice by synthetic wastewater before being introduced into the reactors at a concentration of 5 107 cells per liter. 

Q4. Why is the knowledge of microbial degradation kinetics of 2,6-DCNP unavailable?

Because strain CNP8 is the first bacterium with the ability to degrade 2,6-DCNP, the knowledge of microbial degradation kinetics of 2,6-DCNP is unavailable in the literature. 

Q5. What is the kinetics of the biodegradation of CNP-8?

Because of the inhibition effect of 2,6-DCNP for CNP-8, Haldane’s model (Wang et al., 2010) was used to investigate the kinetics of 2,6-DCNP biodegradation. 

Q6. What is the phylogenetic analysis of the hnp cluster?

The phylogenetic analysis indicated that the hnp cluster originated from the cluster involved in the catabolism of chlorophenols rather than nitrophenols. 

Q7. What is the kinetics of the degradation of 2,6-DCNP?

In summary, although the inhibition effect of 2,6-DCNP against CNP-8 was inevitable, the extremely lower value of Ks (0.03mM) than that of Ki (0.42 mM) indicated that CNP-8 was able to effectively remove 2,6- DCNP from the point of kinetic view. 

Q8. What is the maximum activity of HnpAB for 2,6-DCNP?

Two compounds with GC retention times of 9.87 and 10.74 min,respectively, were detected by the GC-MS analysis of the acetylated products of 2,6-DCNP catalyzed by the purified HnpAB. 

Q9. How much of the degradation rate of 2C4NP was observed after 48 h?

2C4NP was completely removed in the first 48 h; however, 79% (0.238 ± 0.015 mM) of 2C5NP and 88% (0.267 ± 0.024 mM) of 2,6- DCNP remained after this period. 

Q10. What is the kinetics of the two halogenated nitrophenols?

either 2,6-DBNP- or 2,6-DCNP-induced cells of strain CNP-8 have the ability to degrade both halogenated nitrophenols rapidly (Fig. 4). 

Q11. What was the effect of the addition of 2,6-DCNP on the growth of CNP?

for substrate degradation, the specific degradation rates (q) increased with increase of initial 2,6-DCNP concentration, and then declined on further increasing substrate concentration (Fig. S2). 

Q12. What is the role of HnpAB in the catabolism of 2,6-DC?

The enzymatic assay and product identification indicated that HnpAB catalyzed the sequential para- and ortho-position hydroxylation of 2,6-DCNP. 

Q13. What is the kinetics of the degraded nitrophenol?

it degraded (halogenated)-meta-nitrophenol (such as MNP and 2C5NP) via the partial reductive pathways, whereas degraded halogenated para-nitrophenols (such as 2C4NP and 2,6-DBNP) via the oxidative pathways. 

Q14. What was the effect of different initial concentrations of substrate on bacterial growth?

To study the biodegradation kinetics of 2,6-DCNP, the effect of different initial concentrations of substrate (0.05e1.5 mM) on bacterial growth was investigated.