While uncompetitive inhibition necessitates the formation of an enzyme-substrate complex, non-competitive inhibition can occur with or without the presence of the substrate in the reaction.
Uncompetitive inhibition is distinguished from competitive inhibition by two observations: first, uncompetitive inhibition cannot be reversed by increasing [S], and second, as shown in the Lineweaver–Burk plot, parallel lines rather than intersecting lines result from the uncompetitive inhibition experiment. When tertiary amines block the enzyme acetylcholinesterase, this type of behaviour is observed (R3N). Even though such molecules bind to the enzyme in diverse ways, the acyl-intermediate-amine complex cannot be broken down into the enzyme plus the product form of the reaction.
Mechanism
As the inhibitor binds to the ES complex, the amount of ES complex produced decreases. That the ES complex has a lower effective concentration due to the presence of the inhibitor on its surface is explained by the fact that the ES complex is effectively converted to an ESI complex, which is believed to be a different complex altogether. Because it takes longer for the substrate or product to leave the active site as a result of the reduction in the ES complex, the maximal enzyme activity (Vmax) drops as well. Additionally, the decrease in Km – the substrate concentration at which the enzyme can operate at half of its peak velocity, which is frequently used to approximate an enzyme’s affinity for a substrate – may be traced back to the decrease in ES complex. Le Chatelier’s principle stands in opposition to this decline and strives to compensate for the loss of ES by converting more free enzymes to the ES form, resulting in an overall rise in the amount of ES. In general, an increase in ES implies that the enzyme has a high degree of affinity for the substrate being measured. Even though Km is not a perfect predictor of affinity for a substrate because it takes into consideration other aspects as well, an increase in affinity for a substrate will be accompanied by a decrease in Km in most cases.
As a rule, when the substrate concentration is high, uncompetitive inhibition is the most effective. It is not necessary for an uncompetitive inhibitor to be structurally similar to the substrate of the reaction it is blocking. Although the activity of the enzyme will be higher when there is no substrate present, the difference in enzyme activity at low substrate concentrations will be minimal when there is an uncompetitive inhibitor present.
The implications and applications
A range of implications for the impact of uncompetitive inhibition in biological and biochemical systems can be derived from the specific characteristics of uncompetitive inhibition. Uncompetitive inhibition can manifest itself in a variety of different ways in biological systems. In fact, it is frequently discovered that the characteristics of uncompetitive inhibitors that distinguish them from competitive inhibitors, such as their proclivity to perform at their best at high concentrations of substrate, are critical to the normal functioning of various critical body functions.
Participation in the development of cancer processes
Certain kinds of cancer are characterised by the presence of anti-competitive mechanisms. A number of human alkaline phosphatases, including CGAP, have been discovered to be overexpressed in some forms of cancer, and those phosphatases are known to function through uncompetitive inhibition. It has also been shown that amino acids such as leucine and phenylalanine have the ability to block a number of the genes that code for human alkaline phosphatases (TSAPs), which is a novel discovery. Studies of the amino acid residues involved have been carried out in an attempt to regulate alkaline phosphatase activity and learn more about the relevance of this activity to cancer.
Additionally, because it inhibits G6PD, uncompetitive inhibition works in conjunction with TP53 to assist regulate the activity of cancer cells and prevent carcinogenesis in some kinds of the sickness (glucose-6-phosphate dehydrogenase, an enzyme primarily involved in certain metabolic pathways). One of the secondary activities that G6PD is responsible for is assisting in the regulation of reactive oxygen levels, which is important because reactive oxygen species must be kept at appropriate levels in order for cells to survive. When the concentration of G6PD’s substrate is high, uncompetitive inhibition of the enzyme becomes significantly more efficient. Due to an increase in the amount of substrate present, the amount of ES complex present increases as well, and as there is more ES complex available for binding, uncompetitive inhibitors become significantly more active. The mechanism of this inhibition is such that the higher the initial concentration of substrate in the system, the more difficult it is to achieve the maximal velocity of the reaction. Even when starting with low initial substrate concentrations, increasing the concentration of substrate can sometimes be sufficient to restore the enzyme’s function completely or even completely. However, once the initial substrate concentration exceeds a certain point, reaching the maximum enzyme velocity becomes all but impossible. It is likely that uncompetitive inhibition, rather than mixed inhibition, is responsible for the extreme sensitivity to substrate concentration observed within the cancer mechanism. Mixed inhibition, on the other hand, exhibits similar characteristics but is often less sensitive to substrate concentration due to some inhibitor binding to free enzymes regardless of the presence or absence of substrate. Because of their extremely high potency at high substrate concentrations, as well as their overall sensitivity to substrate concentration, uncompetitive inhibitors are the only ones capable of making this type of procedure conceivable.
The role of phospholipids in cell and organelle membranes
Despite the fact that this type of restriction is prevalent in a variety of disorders within biological systems, it is not necessarily exclusive to sickness. It can have a role in a variety of normal body activities. It has been demonstrated that removing lipids from cell membranes and making active sites more accessible through conformational changes can elicit elements resembling the effects of uncompetitive inhibition, indicating that active sites capable of uncompetitive inhibition may be present in membranes (i.e. both KM and VMax decrease). Removing lipids from mitochondrial membranes, in particular, decreases the alpha-helix concentration in mitochondria and causes alterations in the ATPase that are similar to uncompetitive inhibition, according to the findings.
Conclusion
A number of other investigations have also confirmed the presence of uncompetitive enzymes in membranes, which is consistent with this hypothesis. While researching a type of protein known as an Arf protein, which is involved in regulating membrane function, it was discovered that an inhibitor known as BFA trapped one of Arf’s intermediates through the use of uncompetitive inhibitory mechanisms. This demonstrated that this form of inhibition can be found in a variety of cell types and organelles, rather than simply in diseased cells, which was previously thought. The activity of the Golgi apparatus, as well as its involvement in regulating transport across the cell membrane, has been discovered to be correlated with BFA levels in the blood.