Purification to Homogeneity and Characterization of a Novel Pseudomonas putida Chromate Reductase
Open accessAbstract
Cr(VI) (chromate) is a widespread environmental contaminant. Bacterial chromate reductases can convert soluble and toxic chromate to the insoluble and less toxic Cr(III). Bioremediation can therefore be effective in removing chromate from the environment, especially if the bacterial propensity for such removal is enhanced by genetic and biochemical engineering. To clone the chromate reductase-encoding gene, we purified to homogeneity (>600-fold purification) and characterized a novel soluble chromate reductase from Pseudomonas putida , using ammonium sulfate precipitation (55 to 70%), anion-exchange chromatography (DEAE Sepharose CL-6B), chromatofocusing (Polybuffer exchanger 94), and gel filtration (Superose 12 HR 10/30). The enzyme activity was dependent on NADH or NADPH; the temperature and pH optima for chromate reduction were 80°C and 5, respectively; and the K m was 374 μM, with a V max of 1.72 μmol/min/mg of protein. Sulfate inhibited the enzyme activity noncompetitively. The reductase activity remained virtually unaltered after 30 min of exposure to 50°C; even exposure to higher temperatures did not immediately inactivate the enzyme. X-ray absorption near-edge-structure spectra showed quantitative conversion of chromate to Cr(III) during the enzyme reaction.
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Anaerobic conditions are generally required to induce maximum Cr(VI) reduction activity, but some enzyme systems operate under aerobic conditions.
“Anaerobic con-ditions are generally required to induce maximum activity against Cr(VI), but some enzyme systems operate under aerobic conditions, for examples the soluble NAD(P)Hdependent reductases of Pseudomonas ambigua G-1, [61] and Pseudomonas putida [62].”
From Microbial reduction of metals and radionuclides (2003) · cites this paper as [62]
The Abstract describes a soluble Pseudomonas putida chromate reductase dependent on NADH or NADPH, supporting its identification as an example of that enzyme system. Neither the Abstract nor the chromate reductase assays section establishes that it operates under aerobic conditions; the paper also does not substantiate the broader statement that anaerobic conditions are generally required for maximum activity. (LLM, conf=0.82)
Experimental — accuracy not yet measuredCr(VI) bioreduction seems to be initiated by a one-electron transfer from the reductase.
“Thus, Cr(VI) bioreduction seems to be initiated by a one-electron transfer from the reductase.”
From Microbial reduction of metals and radionuclides (2003) · cites this paper as [62]
The Abstract reports that the purified reductase converts Cr(VI) to Cr(III). But the “End product of chromate conversion” section says the assay could not determine whether Cr(V) accumulated, so the paper does not establish that reduction begins with a one-electron transfer. (LLM, conf=0.84)
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Cites
Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4
LAEMMLI, U. K. (1970) · Nature · doi:10.1038/227680a0 · Cited by 249,961 · see citing works
Stohr, J. (1992) · Springer series in surface sciences · doi:10.1007/978-3-662-02853-7 · Cited by 2,313 · see citing works
The complete general secretory pathway in gram-negative bacteria
Pugsley, A. P. (1993) · Microbiological Reviews · doi:10.1128/mr.57.1.50-108.1993 · Cited by 1,761 · see citing works
Heukeshoven, J., & Dernick, R. (1985) · Electrophoresis · doi:10.1002/elps.1150060302 · Cited by 1,348 · see citing works
Reduction of Hexavalent Chromium by Amorphous Iron Sulfide
Patterson, R. R., Fendorf, S. E., & Fendorf, M. (1997) · Environmental Science & Technology · doi:10.1021/es960836v · Cited by 482 · see citing works
Self-citationPacific Northwest Lab., R., Riley, R. G., USDOE, W., et al. (1992) · doi:10.2172/10147081 · Cited by 476 · see citing works
Chromium reduction in Pseudomonas putida
Ishibashi, Y., Cervantes, C., & Silver, S. D. (1990) · Applied and Environmental Microbiology · doi:10.1128/aem.56.7.2268-2270.1990 · Cited by 393 · see citing works
Wang, P.-C., Mori, T., Komori, K., Sasatsu, M., Toda, K., & Ohtake, H. (1989) · Applied and Environmental Microbiology · doi:10.1128/aem.55.7.1665-1669.1989 · Cited by 323 · see citing works
A 13-element Ge detector for fluorescence EXAFS
Cramer, S. P., Tench, O., Yocum, M., et al. (1988) · Nuclear Instruments and Methods in Physics Research Section A Accelerators Spectrometers Detectors and Associated Equipment · doi:10.1016/0168-9002(88)90449-4 · Cited by 253 · see citing works
Suzuki, T., Miyata, N., Horitsu, H., Kawai, K., Takamizawa, K., Tai, Y., & Okazaki, M. (1992) · Journal of Bacteriology · doi:10.1128/jb.174.16.5340-5345.1992 · Cited by 244 · see citing works
Mutagenicity of chromates in bacteria and its relevance to chromate carcinogenesis
Venitt, S., & Levy, L. S. (1974) · Nature · doi:10.1038/250493a0 · Cited by 241 · see citing works
Fate and Transport of Hexavalent Chromium in Undisturbed Heterogeneous Soil
Jardine, P. M., Fendorf, S. E., Mayes, M. A., et al. (1999) · Environmental Science & Technology · doi:10.1021/es981211v · Cited by 230 · see citing works
Self-citationSDS Polyacrylamide Gel Electrophoresis of Proteins
Smith, B. J. (2003) · Humana Press eBooks · doi:10.1385/0-89603-268-x:23 · Cited by 210 · see citing works
Decreased chromate uptake in Pseudomonas fluorescens carrying a chromate resistance plasmid
Ohtake, H., Cervantes, C., & Silver, S. D. (1987) · Journal of Bacteriology · doi:10.1128/jb.169.8.3853-3856.1987 · Cited by 180 · see citing works
DESIGNING MICROORGANISMS FOR THE TREATMENT OF TOXIC WASTES
Timmis, K. N., Steffan, R. J., & Unterman, R. (1994) · Annual Review of Microbiology · doi:10.1146/annurev.mi.48.100194.002521 · Cited by 171 · see citing works
The general secretory pathway in bacteria
Wandersman, C. (1993) · Trends in Microbiology · doi:10.1016/0966-842x(93)90043-q · Cited by 141 · see citing works
Molecular basis of hexavalent chromium carcinogenicity: Effect on gene expression
Wetterhahn, K. E., & Hamilton, J. W. (1989) · The Science of The Total Environment · doi:10.1016/0048-9697(89)90199-x · Cited by 118 · see citing works
Ground-Water Treatment for Chlorinated Solvents
McCarty, P. L., & Semprini, L. (2017) · doi:10.1201/9780203712764-6 · Cited by 107 · see citing works
A carbon starvation survival gene of Pseudomonas putida is regulated by sigma 54
Kim, Y., Watrud, L. S., & Matin, A. (1995) · Journal of Bacteriology · doi:10.1128/jb.177.7.1850-1859.1995 · Cited by 49 · see citing works
Self-citationMatin, A., Little, C., Fraley, C. D., et al. (1995) · Applied and Environmental Microbiology · doi:10.1128/aem.61.9.3323-3328.1995 · Cited by 40 · see citing works
Self-citationMatin, A. (1994) · Annals of the New York Academy of Sciences · doi:10.1111/j.1749-6632.1994.tb47401.x · Cited by 29 · see citing works
Self-citationBajpai, R. K., & Prokop, A. (1994) · New York Academy of Sciences eBooks · Cited by 22 · see citing works
Flagged first, then by citations
Research works printed but not linked: 4
- Greenberg, A. E., J. J. Connors, D. Jenkins, and M. A. Franson (ed.). 1981. Standard methods for the examination of water and wastewater, 15th ed., p. 187-190. American Public Health Association, Washington, D.C.
- Matin, A. 1981. Regulation of enzyme synthesis as studied in continuous culture, p. 69-97. In P. H. Calcott (ed.), Continuous culture of cells, vol. II. CRC Press, Inc., Boca Raton, Fla.
- In J. E. Matthews (ed.), Handbook of bioremediation. Lewis Pub- lishers, Ann Arbor, Mich.
- Norris, R. D. 1994. In-situ bioremediation of soils and groundwater contam- inated with petroleum hydrocarbons, p. 17-37. In J. E. Matthews (ed.), Handbook of bioremediation. Lewis Publishers, Ann Arbor, Mich.
Cited by
Sorted by citations
Microbial reduction of metals and radionuclides
Lloyd, J. R. (2003) · FEMS Microbiology Reviews · doi:10.1016/s0168-6445(03)00044-5 · Cited by 654 · see citing works
Microbial detoxification of metals and radionuclides
Lloyd, J. R., & Lovley, D. R. (2001) · Current Opinion in Biotechnology · doi:10.1016/s0958-1669(00)00207-x · Cited by 342 · see citing works
- Authors
- Park, C. H., Keyhan, M., Wielinga, B., Fendorf, S., & Matin, A.
- Affiliations
- Stanford University (US)
- <!--label omitted: 1-->Department of Microbiology & Immunology1 and
- ORCID
- Venue
- Applied and Environmental Microbiology
- Publisher
- American Society for Microbiology
- Published
- 2000-05
- Volume
- 66
- Issue
- 5
- Pages
- 1788-1795
- ISSN
- 0099-2240 (ISSN-L)
- 1098-5336
- PMC
- PMC101413
- OpenAlex
- W2111298094
- Cited by
- 345
- References
- 22
- Citations by year
- 2026: 9 · 2025: 5 · 2024: 6 · 2023: 10 · 2022: 8 · 2021: 13 · 2020: 18 · 2019: 15 · 2018: 20 · 2017: 11 · 2016: 15 · 2015: 20 · 2014: 14 · 2013: 17 · 2012: 22
- FWCI
- 7.02
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Applied and Environmental Microbiology (publishedVersion)
- PubMed Central (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed (publishedVersion)
- Funders
- U.S. Department of Energy (FG0397ER62494)
- Topics
- Physical Sciences › Environmental Science › Health, Toxicology and Mutagenesis › Chromium effects and bioremediation
- Physical Sciences › Engineering › Biomedical Engineering › Environmental remediation with nanomaterials
- Physical Sciences › Environmental Science › Pollution › Microbial bioremediation and biosurfactants
- Keywords
- chromate reductase, Pseudomonas putida, hexavalent chromium reduction, enzyme purification, enzyme characterization, NADH-dependent reduction, sulfate inhibition, thermal stability, X-ray absorption near edge structure
- MeSH terms
- Biodegradation, Environmental, Chromates, Chromatography, Gel, Chromatography, Ion Exchange, Electrophoresis, Polyacrylamide Gel, Kinetics, Oxidoreductases, Ultracentrifugation, Pseudomonas putida
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