Mutation bacterial

In 1997, Busby and co-workers reported that 2-nitrofluoranthene, an important product of atmospheric transformations (vide infra) was inactive in MCL-5 cells but a potent mutagen in hlAlv2 cells another important atmospheric reaction product, the nitrophenanthrene lactone 2-nitrodibenzopyranone (XI), was inactive in both hlAlv2 and MCL-5 cells. Furthermore, it was nonmutagenic in the forward mutation bacterial assay in the absence of rat liver postmi-tochondrial supernatant (-S9) but was mutagenic with the addition of S9 mix. 

Busby and co-workers (1995) compared the mutagenicities of BaP and five dibenzopyrenes in the Salmonella typhimurium TM677 forward mutation bacterial assay (+ PMS) to three in the MCL-5 human cell assay. The powerful carcinogen dibenzo[a,/]pyrene was 50 times more potent than BaP in human cells (vide supra) however, it was only 1.7 times as potent as BaP in the bacterial assay. Interestingly, there was a 10,000-fold range between the most and least mutagenic PAH in MCL-5 human cells vs a range of only 4 in the bacterial cells. 

In an analogous set of studies at four cities across southern California (see Fig. 10.23), Hannigan and co-workers (1996) employed the Salmonella typhimurium TM677 forward mutation bacterial assay (Skopek et al., 1978b Busby et al., 1994a) to determine the seasonal and spatial variation of the bacterial mutagenicities of fine ambient aerosols. The 1993 annual... 

Working with a mutated bacterial strain, Isbister et al. (62) demonstrated a novel mechanism of aerobic oxidation of dibenzothiophene which involved the specific excision of the sulfur atom from the molecule (Figure 11). Studies with -labeled dibenzothiophene showed the release of the radioactivity into the aqueous phase and ion chromatography showed the appearence of sulfate. There was no radioactive carbon dioxide released when this microorganism was incubated with 14C-labeled dibenzothiophene. GC-MS analysis showed that the oxidation product was 2,2 -dihydroxybiphenyl. Kargi and Robinson (52) also report the release of sulfate from dibenzothiophene. This OSC served as the sole carbon and sulfur source in their cultures of the aerobic thermophile Sulfolobus acidocaldarius. 

Key words Recombineering, Bacteriophage lambda recombination genes, GalK, Point mutation. Bacterial artificial chromosome. Oligonucleotides... 

Mutagenicity. The AJ-nitrosamines, in general, induce mutations in standard bacterial-tester strains (117). As with carcinogenicity, enzymatic activation, typically with Hver microsomal preparations, is required. Certain substituted A/-nitrosamine derivatives (12) induce mutations without microsomal activation (31,33,34). Because the a-acetoxy derivatives can hydroly2e to the corresponding a-hydroxy compounds, this is consistent with the hypothesis that enzymatic oxidation leads to the formation of such unstable a-hydroxy intermediates (13) (118). However, for simple /V-nitrosamines, no systematic relationship has been found between carcinogenicity and mutagenicity (117,119—123). 

A second example is that of an Ala-to-Cys mutation, which causes the fonnation of a rare SH S hydrogen bond between the cysteine and a redox site sulfur and a 50 mV decrease in redox potential (and vice versa) in the bacterial ferredoxins [73]. Here, the side chain contribution of the cysteine is significant however, a backbone shift can also contribute depending on whether the nearby residues allow it to happen. Site-specific mutants have confirmed the redox potential shift [76,77] and the side chain conformation of cysteine but not the backbone shift in the case with crystal structures of both the native and mutant species [78] the latter can be attributed to the specific sequence of the ferre-doxin studied [73]. 

Muragenicuy, genotoxicity Rats, mice, bacterial strains, yeasts Few days Potential to cause mutations, chromosomal damage, and... 

Furthermore, if the antibiotic passes membranes through a specific port of entry, its mutational loss leads to resistance. The lack of the outer membrane protein OprD in P. aeruginosa causes resistance to the (3-lactam antibiotic imipenem. Fosfomycin passes the cytoplasmic membrane via an L-a-glycerol phosphate permease. This transport system is not essential for bacterial growth and therefore mutants with a reduced expression are frequently selected under therapy. 

Figure 4.7 An explanation of the bacterial reverse-mutation test (the Ames test).

In each cycle, the library of mutated genes is first inserted in a standard bacterial host such as Escherichia coli or Bacillus subtilis. Subsequently, bacterial colonies are plated out on agar plates and harvested individually by a colony picker. Each colony is placed in a separate well of a microtiter plate containing nutrient broth, so that the bacteria grow and produce the protein of interest. Because each colony originates... 

Following several cycles of mutagenesis using the E. coli XLl-Red mutator strain and transformation of the plasmid library into E. coli, a total of about 150 000 bacterial colonies were assayed for activity using a colorimetric prescreen [100]. The best mutant Asn336Ser showed a 47-fold increase in activity and a 5.8-fold enhancement... 

Chaudhuri K, Selvaraj S, Pal AK. 1999. Studies on the genotoxicity of endosulfan in bacterial systems. Mutat Res 439 63-67. 

Moriya M, Ohta T, Watanabe K, et al. 1983. Eurther mutagenicity studies on pesticides in bacterial reversion assay systems. Mutat Res 116 185-216. 

Bacterial impermeability Some -lactam antibiotics Mutational loss of porins... 

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