PURIFICATION AND CHARACTERIZATION OF HEAVY METAL INDUCED PROTEASE FROM Pseudomonas fluorescens ATCC 948
Keywords:
Atcc-American Type Culture Collection, Pseudomonas Fluorescens, Ros-Reactive Oxygen Species, Metalloprotease, Environmental PollutionAbstract
The microorganisms that live in association with plant or animal cells rely extensively on the production of extracellular proteases, secondary metabolites and siderophores for their survival. In the gram-negative bacteria, the expression of extracellular products is controlled by the conserved two-component regulatory system consisting sensor kinase GacS and cognate response regulator GacA that also influences the stress tolerance in these bacteria. Oxidative stress is one of the major reasons for the formation of oxidized proteins in aerobic bacteria. During stress, bacteria adapt to the presence of reactive oxygen species (ROS) and oxidation of proteins, by increasing the expression of detoxifying enzymes, protein and DNA repair molecules, which helps in keeping the concentration of these species to sub toxic level by removing the unwanted proteins. This role is attributed to proteases, as they are known to scavenge these oxidized proteins. In response to heavy metal stress (lead, copper and cobalt) in Pseudomonas fluorescens ATCC 948, protease increases in culture supernatant. The highest protease activity was found with exposure of culture to lead. The lead-induced protease was purified to homogeneity and was identified as 33kDa metalloprotease. The purified protease was maximally active at 200C and pH 6.0. The nature of the protease was elucidated using different protease inhibitors, which indicated the protease as metalloprotease. The enzyme is characterized and its role in mechanism of adaptation to heavy metal stress has been discussed.
References
I. Alexandre Bourles et al., (February 2020). Investigating some mechanisms underlying stress metal adaptations of two Burkholderia sensu lato species isolated from New Caledonian ultramafic soils. European Journal of Soil Biology 97.
II. Ali H., Khan E., Llahi I. Environmental chemistry and ecotoxicology of hazardous heavy metals: environmental persistence, toxicity, and bioaccumulation. Hindawi J. Chem. 2019;2019 Article ID 6730305.
III. Alichanidis E and Andrews AT (1977) SoSme properties of the extracellular protease produced by the psychrotrophic bacterium Pseudomonas fluorescens strain AR-11. Biochem. Biophys. Acta., 485: 424-43.
IV. Andersson RE, Hedlung CB,et al.(1979) Thermal inactivation of heat resistant lipase produced by psychrotrophic bacterium Pseudomonas fluorescens. J. Dairy Science., 62: 361-367.
V. Blumer C, Heeb, et al. (1999) Global GacA steered control of cyanide and exoprotease production in Pseudomonas fluorescens involves specific ribosome binding sites. Proc Natl Aca Sci., 96: 14073-14078.
VI. Chopra AK et al. (1985) Purification and characterization of heat-stable proteases from Bacillus stearothermophilus RM-67. J Dairy Sci., 68: 3202-11.
VII. Cornelis P et al (2002) Diversity of siderophore-mediated iron uptake systems in fluorescent pseudomonads: not only pyoverdines. Environ. Microbial. 4:787–798.
VIII. Cornelis P. (2010). Iron uptake and metabolism in pseudomonads. Appl. Microbiol. Biotechnol. 86:1637–1645 10.1007/s00253-010-2550-2
IX. Davies KJ et al (1988) Oxidatively denatured proteins are degraded by an ATP-independent proteolytic pathway in Escherichia coli. Free Radic Biol Med., 5: 225-36.
X. G. Wu, et al A critical review on the bio-removal of hazardous heavy metals from contaminated soils: issues, progress, eco-environmental concerns and opportunities
XI. Gottesman S (1996) Proteases and their targets in E. coli. Annu. Rev. Gene.t, 30: 465-506.
XII. Gottesman S et al (1992) Regulation by proteolysis: energy dependent proteases and their targets. Microbiol. Rev., 56: 592-621.
XIII. Griffith MW, Phillips JD,et al. (1981) Thermostability of proteases and lipases from a number of species of psychrotrophic bacteria of dairy origin. J. Applied Bacteriology., 50: 289-303.
XIV. Grune T, Reinheckel T,et al. (1997) Degradation of oxidized proteins in mammalian cells. FASEB J., 11: 526-34.
XV. Hou WC, Huang DJ, et al. (2002) An aspartic type protease degrades trypsin inhibitors, the major root storage proteins of sweet potato (Ipomoea batatas (L.) Lam cv. Tainong 57). Bot. Bull. Acad.Sin., 43: 271-276.
XVI. Jenal U and Hengge-Aronis R (2003) Regulation by proteolysis in bacterial cells. Curr Opin Microbiol., 6: 163-72.
XVII. Kumura H, Mikawa K, et al. (1993) Purification and some properties of proteinase from Pseudomonas fluorescens No. 33. J Dairy Res., 60: 229-37.
XVIII. Lam MQ et al. (2018) Characterization of detergent compatible protease from halophilic Virgibacillus sp. CD6. 3 Biotech. ;8:104. doi: 10.1007/s13205-018-1133-2.
XIX. Law BA (1979) Reviews of the progress of dairy science: enzymes of psychrotrophic bacteria and their effects on milk and milk products. J. Dairy Research., 46: 573-588.
XX. Lawrence RC (1967) Microbial lipases and esterases. Dairy Science Abstracts., 29: 59-70.
XXI. Mathivanan K et al (2014) Isolation and characterisation of cadmium-resistant bacteria from an industrially polluted coastal ecosystem on the southeast coast of India, Chemistry and Ecology, 30:7, 622-635
XXII. Mitchell SL et al (1989) Properties of heat-stable proteases of Pseudomonas fluorescens: characterization and hydrolysis of milk proteins. J. Dairy Sci., 72: 864-874.
XXIII. Mongkolsuk S and Helmann JD (2002) Regulation of inducible peroxide stress responses. Molecular Microbiology.,45: 9-15.
XXIV. Paik SR, Lee D, et al.(2003) Oxidized glutathione stimulated the amyloid formation of alpha-synuclein. FEBS Lett., 537: 63-7.
XXV. Patel TR, Jackman DM, et al. (1983) Heat-stable protease from Pseudomonas fluorescens T16: purification by affinity column chromatography and characterization. Applied and Environmental Microbiology., 46: 333-7.
XXVI. Riethdorf S, Volker U, et al. (1994) Cloning, nucleotide sequence and expression of Bacillus subtilis lon gene. Journal of Bacteriology., 176: 6518-6527.
XXVII. Rizvi et al (2019) Bioreduction of toxicity influenced by bioactive molecules secreted under metal stress by Azotobacter chroococcum. Ecotoxicology 28(3):302-322.
XXVIII. Sacherer P, Défago G, et al. (1994) Extracellular protease and phospholipase C are controlled by the global regulatory gene gacA in the biocontrol strain Pseudomonas fluorescens CHA0. FEMS Microbiol. Lett., 116: 155-160.
XXIX. Salwan R et al (2019) Trends in extracellular serine proteases of bacteria as detergent bioadditive: alternate and environmental friendly tool for detergent industry. Archives of Microbiology 201(712)
XXX. Sanger F, Nicklen S, et al. (1977) DNA sequencing with chain-terminating inhibitors. Proc. Natl. Acad. Sci., 74: 5463-5467.
XXXI. Singh S, Bajaj BK (2017) Potential application spectrum of microbial proteases for clean and green industrial production. Energ Ecol Environ. 2:370–386.
XXXII. Solanki, P (2021) Microbial proteases: ubiquitous enzymes with innumerable uses. 11(10): 428 Stepaniak L, Fox PF, et al. (1982) Isolation and general characterization of a heat-stable proteinase from Pseudomonas fluorescens AFT 36. Biochem. Biophys. Acta., 717: 376-383.
XXXIII. Tchounwon P (2012).Heavy Metals Toxicity and the Environment 2014 Aug 26:101:133-164.
XXXIV. Turkiewicz M, Gromek E, et al. (1999) Biosynthesis and properties of an extracellular met alloprotease from the antarctic marine bacterium Sphingomonas paucimobilis. J. Biotechnol., 70: 53-60.
XXXV. Whistler CA, Stockwell VO, et al. (2000) Lon protease influence antibiotic production and UV tolerance of Pseudomonas fluorescens Pf-5. Applied and Environmental Microbiology., 66:2718-2725.
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