I want to check the activity of enzymes responsible for heavy metals detoxification can anyone tell me the assay which i should use for the same.
To my best knowledge there are no enzymes for heavy metal detoxifications since they cannot be chemically transformed, they can only be immobilized in certain cell compartments. Metallothioneins and phytochelatines (in plants) are the main mechanisms respnsible for heavy metal immobilization. Perhaps what you are interested in is in the assay of the levels of these polypeptides. I currently do not work on this field but other researchers might inform you on the best commercial or home-made assays.
Alternatively, you might be interested on the enzymatic activity of antioxiant enzymes such as superoxide dismutase, catalase or glutathione peroxidase that detoxify free radicals derived by free heavy metals and induce part of the cell damage attibuted to them. They are actually considered phase II enzymes of xenobiotic detoxification metabolism.
To my best knowledge there are no enzymes for heavy metal detoxifications since they cannot be chemically transformed, they can only be immobilized in certain cell compartments. Metallothioneins and phytochelatines (in plants) are the main mechanisms respnsible for heavy metal immobilization. Perhaps what you are interested in is in the assay of the levels of these polypeptides. I currently do not work on this field but other researchers might inform you on the best commercial or home-made assays.
Alternatively, you might be interested on the enzymatic activity of antioxiant enzymes such as superoxide dismutase, catalase or glutathione peroxidase that detoxify free radicals derived by free heavy metals and induce part of the cell damage attibuted to them. They are actually considered phase II enzymes of xenobiotic detoxification metabolism.
I agree with Myriam. I can't recall any enzyme that is directly involved in heavy metal detox. Organisms mostly sequester heavy metals in complexes.
Actually i want to study the mechanisms involved in detoxification of heavy metals in contaminated soil by microbes......want to explore some new enzymes so thats why i was asking if there is any assay ....
Well then, I suggest you to change the point of view. Metals are elementary species that cannot be detoxified by enzymatic conversion so you should look for microorganisms able to immobilize them... If you have a look at the literature you will find that some microorganisms are able to sequester heavy metals in their walls, while others use scavengers such as polythiols and lead the complexes to lysosomes or vacuoles. For the first ones you can use TEM together with extraction or other physico-chemical techniques. For the second ones, you can assay metallothioneins and phytochelatines.
@ Myriam there are reductases known that convert these metals from one state to other that is less toxic.....i want to explore some other
@Sarabjeet. These reductases are not detoxifying heavy metals, which was your original question. They are just making the metals less toxic to the particular organism being studied.
If you are looking for metal specific reductases, then you should look at methods described in the litterature, like Mohd Shukor suggested.
Mislead?? You really don't need to be an expert in this field to understand that true detoxifying enzymes like SOD/catalase are converting toxic oxygen radicals and H2O2 into non-toxic O2 and H2O, whereas these reductases are making toxic metals less toxic.
I have just seen all the interesting opinions on this topic and I would like to add something. It seems that the differencesmay reside on the language. Some of us find differences between "detoxification" and "toxicity reduction", maybe because our field of expertise includes Xenobiotic Biotransformation Pathways in many organisms from different taxa and even kingdoms. For some of us a metal is not "detoxified" until several biological procesess take place with possible enzymatic redox transformation but necessarly including final conjugation/excretion/immobilization steps as Mohd Shukor. From this point of view, I agree with his/her last message regardign a whole system.
I thought too that maybe the major detoxifying enzymes are oxygenases, in which the assay would be a simple colorimetric or fluorescence assay that measures h2o2.
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