Enzyme inhibitors mercurials

The mechanisms by which metals induce toxic effects or diseases are not well understood. The most toxic heavy metal ions, cadmium, lead, mercury, are potent enzyme inhibitors because the ions are readily polarizable and bind to donor groups in the enzyme, the binding strengths decreasing in the electron-donor element order S > N > O. In-vivo, phosphate and chloride ions are ubiquitous and these may lead to the formation of insoluble species such as lead hydroxophosphate or only slightly soluble mercuric chloride. 

The metallic ions of copper (Cu++), lead (Pb ), and mercury (Hg++) are listed variously as enzyme inhibitors for glutaminolysis. However, not only can they be poisonous to humans, but they can also be poisons or inhibitors for inorganic catalysts. 

Many chemicals can bind to enzymes and either eliminate or drastically reduce their catalytic ability. These chemicals, called enzyme inhibitors, have been used for hundreds of years. When she poisoned her victims with arsenic, Lucretia Borgia was unaware that it was binding to the thiol groups of cysteine amino acids in the proteins of her victims and thus interfering with the formation of disulfide bonds needed to stabilize the tertiary structure of enzymes. However, she was well aware of the deadly toxicity of heavy metal salts like arsenic and mercury. When you take penicillin for a bacterial infection, you are taking another enzyme inhibitor. Penicillin inhibits several enzymes that are involved in the synthesis of bacterial cell walls. 

The activity of an enzyme is destroyed if some molecule other than the substrate specific to that enzyme binds to the active site and blocks entry of the substrate. Such substances are called enzyme inhibitors. Nerve poisons and certain toxic metal ions, such as lead and mercury, are believed to act in this way to inhibit enzyme activity. Some other poisons act by attaching elsewhere on the enzyme, thereby distorting the active site so that the substrate no longer fits. 

Inorganic and organic mercury compounds have a strong affinity for thiol chemical groups. Most proteins and aU enzymes contain these groups so that mercury readily is bound to body tissues. Most mercury compounds are potent enzyme inhibitors which affects membrane permeability, which in turn affects nerve conduction and tissue respiration. 

Enzyme inhibitors also need to be taken into account. These include heavy metal ions, such as silvei mercury, and sulfydryl-reacting organic compounds, such as p-chloromercuribenzoate (due to their reaction with the free S-H groups at the active site of many enzymes, especially the oxidases). 

The fact that ATP and CTP bind to the same site follows from the observation that adding ATP to the inhibited enzyme by CTP reduces or reverses the inhibition, presumably because ATP competes with CTP for the same site. The fact that CTP binds to an allosteric site (i.e., it is not a competitive inhibitor) follows from the so-called desensitization effect. Addition of mercurials [e.g., p-mercuribenzoate (PMB)] reduces or eliminates the inhibition by CTP. However, it has no effect on the enzymatic activity of ATCase, presumably because the mercurials affect the regulatory subunits but not the catalytic site. As for the mechanism of cooperativity (both positive and negative), it is known that CTP does induce changes in the quaternary structure of the enzyme. 

The buried Cys-212 of human carbonic anhydrase B (3 pM) is virtually unreactive towards 2-chloromercuric-4-nitrophenol (60 pM) at pH 9.2, but upon the addition of only 40 pM CN , the half-life drops to 10 minutes which is an, at least, 75-fold rate enhancement. On first analysis, this would suggest that inhibitor binding to the enzyme has produced a conformational change or altered the — SH environment of the Cys—212. This is unexpected. How would you prove by kinetic experiments that the CN is binding to the mercury compound and not the enzyme and that this is changing the reactivity. The rate reaches a constant value at high [CN ]. 

There are several such toxic agents that cause considerable medical, public and political concern. Two examples are discussed here the heavy metal ions (e.g. lead, mercury, copper, cadmium) and the fluorophosphonates. Heavy metal ions readily form complexes with organic compounds which are lipid soluble so that they readily enter cells, where the ions bind to amino acid groups in the active site of enzymes. These two types of inhibitors are discussed in Boxes 3.5 and 3.6. There is also concern that some chemicals in the environment, (e.g. those found in industrial effluents, rubbish tips and agricultural sprays), although present at very low levels, can react with enhanced reactivity groups in enzymes. Consequently, only minute amounts concentrations are effective inhibitors and therefore can be toxic. It is suggested that they are responsible for some non-specific or even specific diseases (e.g. breast tumours). 

There are a number of metals that are irreversible inhibitors of enzymes. Those that are found in increasing quantities in the environment and hence are causing concern include lead, mercury, cadmium and copper. Enzymes that possess the amino acid cysteine in their active site are most vulnerable, since the sulphydryl group (-SH) in this site readily reacts with the metal ions (Appendix 3.7). 

Examples of noncompetitive inhibitors are the heavy metals which act as poisons such as lead, mercury and silver ions. Mercury is attracted to -SH(thiol) groups in the active sites of enzymes... 

The principle of combination of electrochemical glucose oxidase biosensor with the clean-up method for direct extraction and determination of methyl mercury has been successfully demonstrated. The extraction of methyl mercury from the organic solvent has been based on invertase enzyme inhibition. The combination of very low concentration of invertase enzyme and 10 min of incubation time allows the detection of methyl mercury at 5 ppb level. Our method permits the detection of this inhibitor below the legal limit given by the European Union with good recoveries when fish samples were measured. 

In-vitro approach Data are available in abundance concerning metal effects on isolated chloroplasts (for a review, see Clijsters and Van Assche, 1985). All the metals studied were found to be potential inhibitors of photosystem 2 (PS 2) photosystem 1 (PS 1) was reported to be less sensitive. From the in-vitro experiments, at least two potential metal-sensitive sites can be derived in the photosynthetic electron transport chain the water-splitting enzyme at the oxidising side of PS 2, and the NADPH-oxido-reductase (an enzyme with functional SH-groups) at the reducing side of PS 1 (Clijsters and Van Assche, 1985). Moreover, in vitro, non cyclic photophosphorylation was very sensitive to lead (Hampp et al., 1973 b) and mercury (Honeycutt and Korgmann, 1972). Both cyclic and non-cyclic photophosphorylation were proven to be inhibited by excess of copper (Uribe and Stark, 1982) and cadmium (Lucero et al, 1976). 

Inhibition. Since papain, ficin, and bromelain are all enzymes whose activity depends on a free SH group, it is to be expected that all thiol reagents act as inhibitors. Thus, a-halogen acids or amides and N-ethyl-maleimide irreversibly inhibit the thiol proteases. Heavy metal ions and organic mercurial salts inhibit in a fashion that can be reversed by low molecular weight thiols, particularly in the presence of EDTA which... 

It has been noted that certain substances slow down and even stop enzyme-controlled reactions they are called inhibitors. Competitive inhibition occurs when a molecule has a similar molecular configuration to the substrate it therefore competes with the normal substrate for the active site and may slow down the reaction. The degree of inhibition depends on the relative concentrations of the substrate and inhibitor. Non-competitive inhibition occurs when the inhibitor attaches itself permanently to the active site the extent of inhibition depends entirely on the inhibitor concentration and cannot be altered by changing the amount of substrate present. Arsenic and heavy metals such as mercury and silver are toxic because they are inhibitors (non-competitive). Nerve gas, developed during the Second World War, is another example of an inhibitor it combines competitively with the enzyme cholinesterase and slows down the transmission of nerve impulses from one cell to another. 

Mercury is a reactive element and its toxicity is probably due to interaction with proteins. Mercury has a particular affinity for sulphydryl groups in proteins and consequently is an inhibitor of various enzymes such as membrane ATPase, which are sulphydryl dependent. It can also react with amino, phosphoryl and carboxyl groups. Brain pyruvate metabolism is known to be inhibited by mercury, as are lactate dehydrogenase and fatty acid synthetase. The accumulation of mercury in lysosomes increases the activity of lysomal acid phosphatase which may be a cause of toxicity as lysosomal damage releases various hydrolytic enzymes into the cell, which can then cause cellular damage. Mercury accumulates in the kidney and is believed to cause uncoupling of oxidative phsophorylation in the mitochondria of the kidney cells. Thus, a number of mitochondrial enzymes are inhibited by Hg2+. These effects on the mitochondria will lead to a reduction of respiratory control in the renal cells and their functions such as solute reabsorption, will be compromised. 

Noncompetitive inhibition occurs when the inhibition depends only on the concentration of the inhibitor. This is usually caused by adsorption of the inhibitor at a site other than the active site but one which is necessary for activation. In other words, an inactive derivative of the enzyme is formed. Examples are the reaction of the heavy metals mercury, silver, and lead with sulfhydryl groups (—SH) on the enzyme. The sulfhydryl group is tied up by the heavy metal (ESH + Ag" " —> ESAg + H" ), and this reaction is irreversible. This is why heavy metals are poisons they inactivate enzymes in the body. 

As ATP is often the substrate in the case of enzyme-substrate-metal complexes, most metals are active for they mostly bind to the triphosphate. Copper (II), mercury (II), and other very strong Lewis acceptors are inhibitors as they bind to the ring nitrogens of ATP and in enzymes they could also block essential sulfhydryls. 

---