Neither the Kastle-Meyer test nor the luminol test can identify whose blood is involved, but once a stain has been determined to be blood, traces of DNA can be extracted and an identification carried out. In the example of the jeans, DNA analysis was able to exclude the blood coming from the owner of the jeans. Luminol analysis does have drawbacks. Its chemiluminescence can also be triggered by a number of substances such as copper-containing compounds and bleaching agents.
Had the jeans been washed with a detergent containing a bleaching agent, the blood would not have been detected. Criminals aware of this have been known to try to wash away traces of their crime with bleach. The result is that residual bleach makes the entire crime scene produce the typical blue glow, which effectively camouflages any blood stain.
And if you want to see a really impressive glow, spray a piece of liver with a luminol test solution. Enter your keywords. Sign-Up Here. How does that happen? A foam forms when bubbles of a gas are trapped in a liquid or solid. In this case oxygen is generated when hydrogen peroxide breaks down into oxygen and water on contact with catalase, an enzyme found in liver. Is it true that the Beatles wrote a song about LSD? Is it true that you cannot eat polar bear liver?
What is Guarana? Furthermore, Stern showed that the hemin group of the enzyme can react with compounds such as cyanides, sulfides, fluorides and he also showed that the enzyme active group had a ferric complex identical to the protoporphyrin found in the hemoglobin of red blood cells Stern, Thereafter, Sumner and Dounce purified and crystallized bovine catalase. Modified from Sies In the s, it was elucidated the role of key residues in the active site of the enzyme, such as the distal histidine, and their importance for the stabilization of the tertiary structure was discussed by Nakatani In , catalase Compound I was identified in intact eukaryotic cells, proving the existence of H 2 O 2 in normal aerobic metabolism Sies and Chance, Kirkman and Gaetani reported that NADPH was the enzyme cofactor bound to catalase , which was further confirmed following X-ray analysis of catalase structures Fita and Rossmann, a.
Finally, recombinant phage clones containing the human catalase gene were isolated and characterized Quan et al. In this context, Nenoi et al. Recently, Glorieux et al. This novel regulatory mechanism is involved during cancer cells adaptation to chronic exposure to H 2 O 2 and may have therapeutic consequences for various diseases and metabolic disorders.
With the increasing number of complete sequences available, distinct homologies were detected and catalases came to be classified in three groups based on their structure and function. The first and the second group contain heme-containing enzymes, namely typical or true catalases and catalase-peroxidases, whereas the third group contains non-heme manganese catalases Zamocky and Koller, ; Zamocky et al.
The members of this largest group are found in aerobically respiring organisms. In contrast, in anaerobic bacteria, catalases proteins are generally not expressed. Most of these catalases are homotetramers, between and kDa in size and contain four prosthetic groups.
In the majority of true catalases, a ferric protoporphyrin IX was found in the active center namely heme b , similar to the prosthetic group of human hemoglobin. Following phylogenetic analyses the typical catalases can also be divided into three main clades.
Clade 1 contains bacterial, algal and plant catalases with small-subunit size 55—69 kDa using heme b as the prosthetic group.
Clade 3 is the most abundant subfamily; catalases from this subgroup are found in archaebacterial, fungi, protists, plants and animals. The human catalase belongs to this clade and is characterized by a small subunit 62 kDa , with heme b as its prosthetic group and NADPH as cofactor. These proteins have been found in fungi, archeobacteria and bacteria.
Their molecular weight varies between and kDa and they are generally homodimers. The catalase activity degrading hydrogen peroxide is less efficient than in typical catalases but catalase-peroxidases have a better affinity for their substrate H 2 O 2.
Catalase-peroxidases are also significantly more sensitive than typical catalases to inactivation by pH and temperature. The well-known horseradish peroxidase, currently employed in immunoblotting experiments, is one example of catalase-peroxidase.
These enzymes have been found exclusively in bacteria. Manganese catalases utilize two manganese ions in the active site, they can form oligomeric structures measuring between and kDa and have no significant homology with either typical catalases or catalase-peroxidases. The catalytic reaction is completely different to other types of catalases. Like typical catalases, the catalase reaction occurs in two-step. Human catalase contains four identical subunits of 62 kDa, each subunit containing four distinct domains and one prosthetic heme group Nagem et al.
Although amino acid sequences do not have high identities between all typical catalases, the tridimensional structure is highly conserved.
The investigation of inhibitory mechanisms by which cyanide and 3-amino-1,2,4-triazole ATA inhibit human catalase allowed to understand the enzyme activity Putnam et al. Cyanide nitrogen blocks heme access to other potential ligands. It interacts with the distal histidine and an asparagine residue suggesting that it competes with hydrogen peroxide for heme binding. Meanwhile, ATA interacts with the distal histidine leading to an adduct formation and thereby blocks the catalase reaction.
During the enzymatic reaction leading to H 2 O 2 destruction, catalase is first oxidized to a hypervalent iron intermediate, known as compound I Cpd I , which is then reduced back to the resting state by a second H 2 O 2 molecule.
The first reaction is characterized by the oxidation of the heme protein by a single H 2 O 2 molecule leading to the formation of Cpd I , an oxoferryl porphyrin cation radical Jones and Dunford, This second reaction is particularly efficient in some catalases compared to other heme proteins such as myoglobin Matsui et al. Labeling studies have shown that both H 2 O and O 2 molecules are formed from the same molecule of H 2 O 2 Vlasits et al.
Fita and Rossmann b proposed that two molecules of H 2 O 2 were sequentially transferred to the oxoferryl group of Cpd I , where the distal histidine residue plays a role of acid-base catalyst.
Although mutation of distal histidine suppresses the ability to form Cpd I Nakatani, , some authors claimed that reaction occurs as a direct mechanism and histidine does not play a crucial role in catalysis Kato et al.
In the presence of one-electron donors such as phenols, ferrocyanide, salicylic acid, NO, superoxide anions and low H 2 O 2 concentrations, Cpd I may undergo a one-electron reduction towards the inactive compound II Cpd II intermediate, which transforms back to the resting state through another one-electron reduction step reviewed in Bauer, In the presence of another one-electron donor, Cpd II will return to a resting state.
In this intermediate state, iron is at an oxyferrous state O 2 -Fe II -heme. Then, Cpd III goes back to a resting state or leads to the inactivation of the catalase. Regarding catalase localization within the cell, it should be noted that catalase is mainly located in peroxisomes because it contains a sequence signal recognized by some peroxisome receptors.
Contrary to mitochondria, proteins located within peroxisomes are all of nuclear origin and should be imported. Indeed, it is generally accepted that catalase monomers are imported into peroxisomes where tetramerization and heme addition occurs Lazarow and De Duve, The apoprotein monomer enters into the peroxisome by a peroxisome-targeting signal sequence PTS present on the carboxy-terminal tail of catalase.
The most common targeting signal is the SKL serine-lysine-leucine but catalase is characterized by a different signal, namely the KANL lysine-alanine-asparagine-leucine signal Purdue and Lazarow, Some diseases related to defects in peroxisome biogenesis, such as Zellweger syndrome, are characterized by mutations in the PEX5p receptor and cellular H 2 O 2 overproduction due to a catalase default import in peroxisomes Wanders et al.
It has been shown that overexpressing receptor mutants in PEX5-deficient CHO cells, drastically reduce the import of proteins such as catalase Shimozawa et al. Some authors have observed that catalase with the SKL signal and not KANL had a better import capacity and could be transported into the peroxisome even in the case of mutations in the PTS1 receptor Koepke et al.
It has been shown that PEX5p recognizes catalase which is already folded and interacts with other receptors such as PEX13p for proper import into the peroxisome Otera and Fujiki, Studies are underway to validate, through clinical trials, whether catalase SKL has a therapeutic potential for diseases with peroxisome biogenesis disorders.
Recently, it has been reported that PEX19p, an essential protein for peroxisome biogenesis, interacts with Valosin-containing protein VCP and regulates the catalase cytoplasmic localization, a potential feedback mechanism modulating H 2 O 2 levels Murakami et al.
Interestingly, the existence of a cytosolic catalase, in its active tetrameric conformation, has been reported Middelkoop et al. Indeed, catalase may bind cytosolic proteins such as Grb2 and SHP2, to protect them from potential oxidative damage Yano et al.
These proteins are linked to the membrane by a pleckstrin homology PH domain, so it is common to find catalase in fractions including membrane proteins and membrane-associated proteins. In this context, it has been shown that catalase can be localized at the cytoplasmic membrane, specifically at the surface of cancer cells Bauer, Furthermore, localized expression of catalase on the membrane of tumor cells is not in disagreement with the finding of lower total catalase concentration in malignant cells, as the membrane comprises a minority of the total cellular material.
This soluble extracellular catalase was protective for the tumor cells. Finally, catalase has been also localized in the mitochondria of rat cardiomyocytes Radi et al. The human catalase is expressed in every organ and the highest levels of activity are measured in the liver, kidney and red blood cells Winternitz and Meloy, The first function assigned to catalase is the dismutation of H 2 O 2 into oxygen and water without consummation of endogenous reducing equivalents, an important role in cell defense against oxidative damage by H 2 O 2.
To note that H 2 O 2 is not only toxic by its ability to form other ROS, like hydroxyl radical through the Fenton reaction Fenton, but as was nicely recently reviewed by Sies, H 2 O 2 acting as a second messenger is involved in many biological processes including changes of morphology, proliferation, signaling i.
It has also been reported that catalase may decompose peroxynitrite Gebicka and Didik, ; Heinzelmann and Bauer, , oxidize nitric oxide to nitrite Wink and Mitchell, ; Brunelli et al. Catalase also exhibits low oxidase activity O 2 -dependent oxidation of organic substrates Vetrano et al. Thus, catalase may also have additional roles such as the detoxification or activation of toxic and anti-tumor compounds.
For instance, catalase has been detected in mouse oocytes most likely to protect the genome from oxidative damage during meiotic maturation Park et al. Within this framework, several studies have shown a change in catalase expression in cancer cells became resistant to chemotherapies Akman et al. Thus, a potential role of catalase during the acquisition of cancer cell resistance to chemotherapeutic agents was explored by overexpressing the human enzyme in MCF-7 cells, a human derived breast cancer cell line Glorieux et al.
No particular resistance against conventional chemotherapies like doxorubicin, cisplatin and paclitaxel was observed in cells overexpressing catalase but they were more resistant to the pro-oxidant effect induced by an H 2 O 2 -generating system Glorieux et al. In such animals, the development of mitochondrial deletions was reduced and heart disease and the onset of cataracts were delayed. Regarding catalase down-regulation, no particular sensitivity was observed as catalase-deficient mice are viable and fertile Ho et al.
They develop normally with a normal hematological profile, but after trauma the mitochondria shows defects in the oxidative phosphorylation. Note that humans may also be deficient in catalase, a condition known as acatalasemia that is characterized by a low catalase rate, but it is still rare and usually benign Goth et al. In this context, there are benign polymorphisms of the catalase gene for which no change of catalase expression or activity was detected Goth et al.
To note that the catalase gene encodes one single protein of amino acids and the single locus has been mapped to chromosome 11p13 Wieacker et al. The length of the catalase gene is 34 kb, it contains 12 introns and 13 exons generating a mRNA of bp Quan et al. Conversely, some catalase mutations provoke changes in either catalase expression or activity and may be associated with some diseases.
Recent studies have focused on the associations of catalase polymorphisms with various types of cancer but many inconsistent results about the relationship between the catalase gene polymorphism and cancer risk were reported.
Recently, two meta-analyses pointed out a correlation exists between this polymorphism CT and prostate cancer Liu et al. Different catalase mutations in patients can cause decreased catalase activity leading to increased H 2 O 2 concentrations in the blood and tissues. Depending on the mutations, such patients may be subject to an increased risk of type 2 diabetes, vitiligo and increased blood pressure Goth et al.
When the mutations are located in exons as in Japanese and Hungarian acatalasemia, a truncated or mutated catalase is synthetized and functionally less active. This table was adapted from Goth et al. Decreased activity of catalase has also been observed in various genetic alterations, for example, loss of alleles i.
It is generally accepted that the cellular maintenance of redox homeostasis is controlled by a complex network of antioxidant enzymes i. Nevertheless, the molecular mechanisms regulating the expression of catalase — the oldest known and first discovered antioxidant enzyme — are independent of this pathway and not totally elucidated.
Therefore, the fine-tuning regulation of this enzyme should be prior elucidated in order to find a new approach to modulate the antioxidant status in cancer cells. Interestingly, despite the existence of diverse protection mechanisms against oxidant injuries, a consensus emerged in the scientific literature about an alteration of redox homeostasis within tumor cells. Indeed, they produce large amounts of ROS that are involved in the maintenance of genetic instability favoring cancer cell proliferation.
Meanwhile, altered expression levels of catalase have been reported in cancer tissues as compared to their normal counterparts. Thus, as compared to normal tissues of the same origin, some authors reported an increased catalase expression in tumors Sander et al. For instance, we have reported an important decrease of catalase activity in different cancer cell lines, as shown in Table 2 Verrax et al.
Briefly, catalase levels can vary after short treatments to H 2 O 2 Rohrdanz and Kahl, ; Rohrdanz et al. Although mechanisms controlling catalase expression have been partially elucidated, the decreased catalase expression in cancer cells still remains an unanswered question.
Data were analyzed by unpaired t -test. The regulation of catalase expression in cancer cells is a complex process because different levels of regulation are thought to be involved. In a recent review, we discussed the different mechanisms playing a potential role in the regulation of its expression in both healthy and tumor cells Glorieux et al.
They include transcriptional regulation, represented by the activity of transcription factors that induce or repress the transcriptional activity of catalase promoters, post-transcriptional regulation mRNA stability and post-translational modification phosphorylation and ubiquitination of the protein.
In addition, epigenetic DNA methylation, modifications of histones changes or genetic alterations can also be involved playing a role in governing proper levels of catalase activity in these cells. Regarding transcription it should be noted that the catalase gene has all the characteristics of a housekeeping gene no TATA box, no INR sequence, high GC content in promoter and a core promoter which is highly conserved among species Quan et al.
In this core promoter, the presence of DNA binding sites for transcription factors like NF-Y and Sp1 has an essential role in the positive regulation of catalase expression Nenoi et al.
Additional transcription factors have also been involved in this regulatory process. Specifically, we investigated the transcriptional regulatory mechanism controlling catalase expression in human mammary cell lines. To this end, we have made a human breast MCF-7 cancer cell line resistant to oxidative stress, the so-called Resox cells. These cells show decreased ROS basal levels and an increased activity of some antioxidant enzymes, notably catalase Dejeans et al. Thus, cancer adaptation to oxidative stress, regulated by transcriptional factors through chromatin remodeling appears as a new mechanism to target cancer cells.
These cells show increased expression of catalase. HAT: Histobe acetyltransferase. Adapted from Glorieux et al. As previously mentioned, cancer cells are generally deficient in antioxidant enzymes, thus, any increase in ROS levels would be a menace to the precarious redox balance of cancer cells making them vulnerable to an additional oxidative stress.
Given that weakness, the loss of redox homeostasis represents an interesting target for research and development of new molecules with antitumor activity and numerous drugs are currently being clinically evaluated.
Therefore, several strategies have been developed looking for the disruption of tumor cell redox homeostasis and a subsequent cancer cell death Demizu et al.
One of these strategies is the use of ROS-generating compounds such as arsenic trioxide ATO , currently employed against promyelocytic leukemia Valenzuela et al. Indeed, the impairment of mitochondrial function due to increased levels of superoxide anion is supposed to be the main mechanism of both chemotherapeutic drugs Thayer, ; Pelicano et al.
A decrease in antioxidant levels has also been proposed. The development of specific inhibitors of thioredoxin and thioredoxin reductases was also carried out. Inhibitors of thioredoxin, such as PX, were shown to have potent antitumor activities Welsh et al.
Conversely, the overexpression of Trx1 is correlated with resistance to anti-cancer drugs Baker et al. As quinones display redox cycling abilities thus generating ROS Kappus and Sies, , the association of menadione a naphthoquinone derivative and ascorbate Figure 3 , was employed to trigger tumor cell death Verrax et al. We hypothesized that H 2 O 2 , issued from the redox cycling, is the oxidant species responsible for antitumor effects observed both in vitro and in vivo Verrax et al.
The induced oxidative stress provokes cell necrosis by a wide variety of processes including ATP depletion Verrax et al. Based on the vulnerability of tumor cells to an oxidative stress, we have induced the alteration of their intracellular redox homeostasis as a new strategy in the research and development of new antitumor drugs Benites et al.
A similar approach has been recently developed by using the SnFe 2 O 4 nanocrystals, a heterogeneous Fenton catalyst, which once internalized into the cancer cells, convert H 2 O 2 into hydroxyl radicals inducing apoptotic cell death. In normal cells, the oxidative injury induced by SnFe 2 O 4 is prevented by catalase Lee et al.
The biological activity showed by menadione mainly relies upon its ability to accept one electron from ascorbate to form a semiquinone radical. When the semiquinone is reduced back to its quinone form, superoxide anion is produced which by dismutation generates hydrogen peroxide.
Despite that Resox cells display high antioxidant defenses Dejeans et al. Arsenic trioxide ATO decreases catalase expression in breast cancer cells and sensitizes them to pro-oxidant drugs. Under stress conditions, the antioxidant enzyme catalase plays a major role by detoxifying H 2 O 2.
As consequences, a change of its activity or expression will lead to pathological processes as Zellweger syndrome, acatalasemia or WAGR syndrome. The subcellular localization of catalase is mainly peroxisomal but a shuttle between this organelle and cytoplasm exists and may be involved in the protection of key cellular elements i.
Our group and others demonstrated that catalase expression is also altered in cancer cells, most likely to favor cell proliferation by inducing genetic instability and activation of oncogenes. The regulation of catalase expression appears to be mainly controlled at transcriptional levels although other mechanisms may also be involved.
Therefore, catalase can be a future therapeutic target in the context of cancer by using pro-oxidant approaches. The authors thank Professor Helmut Sies for the splendid discussion and his precious input. Akca, H. Enzyme Inhib. Search in Google Scholar. Akman, S. Antioxidant and xenobiotic-metabolizing enzyme gene expression in doxorubicin-resistant MCF-7 breast cancer cells.
Cancer Res. Arenas, P. Echo-friendly synthesis and antiproliferative evaluation of oxygen substituted diaryl ketones.
Molecules 18 , — Baker, A. Expression of antioxidant enzymes in human prostatic adenocarcinoma. Prostate 32 , — The antitumor thioredoxin-1 inhibitor PX 1-methylpropyl 2-imidazolyl disulfide decreases thioredoxin-1 and VEGF levels in cancer patient plasma.
J Lab Clin Med. Barletta, C. Tumori 71 , — Bauer, G. Tumor cell-protective catalase as a novel target for rational therapeutic approaches based on specific intercellular ROS signaling. Anticancer Res. Increasing the endogenous NO level causes catalase inactivation and reactivation of intercellular apoptosis signaling specifically in tumor cells.
Redox Biol. Mechanisms of selective antitumor action of cold atmospheric plasma-derived reactive oxygen and nitrogen species. Plasma Process Polym. The antitumor effect of single-domain antibodies directed towards membrane-associated catalase and superoxide dismutase. Beck, R. Hsp90 cleavage by an oxidative stress leads to Bcr-Abl degradation and leukemia cell death. What is the color and consistency of this mixture? Put one drop of the mixture on a clean part of the large plate and add one drop of hydrogen peroxide to it.
Compared with the untreated blended liver, did more, less or about the same amount of bubbles form? Did they form more slowly, more quickly or at about the same rate?
Did more, less or about the same amount of bubbles form? Cover the bowl and microwave it on high for 20 seconds. How does the blended liver look after heating? Remove a drop-size amount of the heated liver and put it on a clean part of the large plate. Add one drop of hydrogen peroxide to it. Which condition s makes it work the worst? Why do you think this is so?
For example, try freezing some blended liver or mixing it with salt and then test the enzyme's activity. Or you could try adding more than one teaspoon of vinegar or baking soda and then test the enzyme. Under which conditions does the enzyme work well, and under which ones does it work poorly?
One protein that is fun to digest using bromelain is gelatin, which is found in many puddings and gelatinous desserts. How do different conditions affect the ability of bromelain to digest proteins? Observations and results. When exposed to hydrogen peroxide, did the blended liver bubble less when mixed with either the vinegar or baking soda compared with when it was untreated? Did it bubble even less after it was microwaved? An enzyme needs certain conditions to work, and the ideal environment can be a hint as to where the enzyme normally works in the body.
And because different body tissues have distinct environments—acidic or warm—each enzyme is tuned to work best under specific conditions. Different tissues in the body have different pHs pH is a measure of how basic or acidic a solution is. The liver maintains a neutral pH about pH 7 , which is easiest for its enzymes, such as catalase, to work in.
Consequently, when exposed to hydrogen peroxide the liver should have produced more bubbles oxygen gas , and at a faster rate, when it was untreated than when exposed to vinegar or baking soda.
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