Proteases (proteinases, peptidases and proteolytic enzymes) play a very important role in human life. To date, more than 500 of these enzymes have been identified in the human body, which are encoded by 2% of all genes. Proteolytic activity is observed in all life forms and viruses.
Main classification
Based on the catalytic residue, proteases can be divided into 7 broad groups:
- Serine - use of serine alcohol.
- Cysteine - the use of cysteine thiol.
- Threonine - turnover of secondary alcohol threonine.
- Aspartic - using aspartate carboxylic acid.
- Glutamic - using the carboxylic acid glutamate.
- Metalloprotease - metal turnover, usually zinc.
- Asparagine peptide lyases - Asparagine is used to perform an exclusion reaction (not requiring water).
Proteases were first grouped into 84 families according to their proteolytic activity, andclassified under 4 catalytic types:
- serine;
- cysteine;
- aspartic;
- metal.
Meaning
Total proteolytic activity plays an important role in many body processes. These are fertilization, digestion, growth, maturation, aging and even death. Proteases regulate many physiological processes by controlling the activation of synthesis and the degradation of protein foods. They play an important role in the reproduction and spread of viruses, bacteria and parasites and are therefore responsible for the efficient transmission of diseases caused by pathogenic substances. These enzymes allow tumor cells to divide, fill the intercellular space and blood vessels of the lungs, and spread to other tissues of the body.
Biological functions
Proteolytic activity has the following functions:
- Post-translational processing. Includes removal of methionine and/or conversion of an inactive or non-functional protein to an active unit.
- Cleavage of precursor proteins. This is necessary to ensure that the protease is only activated in the correct location and context. Improper proteolytic activity can be very damaging to the body.
- Protein degradation. May occur intracellularly or extracellularly. Performs a number of functions: removes damaged and abnormal proteins; prevents their accumulation; serves to regulate cellular processes by removingenzymes.
- Digestion. Proteins obtained from food are broken down into peptide chains by pepsin, trypsin, chymotrypsin and elastase. To prevent inappropriate or premature activation of digestive enzymes (which can cause pancreatitis), they act as an inactive zymogen.
Enzymes
Proteolytic enzymes are found in bacteria, viruses, some types of algae and plants. But most of them are in animals. There are various types of proteolytic activity of enzymes. They are classified according to the sites in which the breakdown of proteins is catalyzed. The two main groups are exopeptides and endopeptidases. Within the body, protein materials are initially attacked by pepsin. When the protein is passed into the small intestine, it is partially digested by the stomach. Here it is exposed to proteolytic enzymes secreted by the pancreas. Then pancreatic enzymes are activated in the intestine, converting proteins into amino acids that are easily absorbed by its walls. Thus, the pancreas is protected from self-digestion.
Bacteria
Microbial proteases are one of the important groups in the industrial-commercial production of enzymes. Studies have been conducted to determine the proteolytic activity of bacteria in order to elucidate their roles within the pathogenesis of infectious diseases. The focus was on the examination of lactic acid bacteria from various yoghurts and fermented milks. They are widely distributed in nature. These are lactobacilli, lactococci, bifidobacteria, streptococci, enterococci and sporolactobacilli. They are divided into species, subspecies, variants and strains.
Proteolytic activity is a very important characteristic of lactic acid bacteria. Bacterial proteases are enzymes that catalyze hydrolysis peptide bonds in proteins and polypeptides. They play a significant role in industrial biotechnology and pharmaceuticals. Studies have shown that 13 strains have proteolytic activity. Five of them, namely L1, L2, L6, L7, L9 showed the highest activity.
Peptins
Proteolytic activity of pepsin is measured under the influence of a magnetic field on the body. The molecular structure of pepsin is characterized by D-spatial symmetry. The inactive proenzyme pepsinogen is synthesized within the cells of the gastric mucosa. It is also present in various biological fluids (blood, urine, seminal and cerebrospinal fluid). Pepsinogen is characterized by autocatalytic activation. Its secretion is stimulated by the vagus nerve, sympathetic fibers, gastrin, histamine, secretin, and cholecystokinin. Gastrin acts as a parietal cell stimulator. This polypeptide exists in 2 forms containing 34 and 17 amino acids. Measurements of the proteolytic activity of pepsin in relation to standard hemoglobin revealed similar changes in the digestive activity of the enzyme.
Proteolysis and disease
Anomalous proteolyticactivity is associated with many diseases. In pancreatitis, leakage of proteases and their premature activation in the pancreas leads to auto-ignition of the pancreas. People with diabetes may have an increased activity of lysosomes, and the degradation of some proteins may increase significantly. Chronic inflammatory diseases (rheumatoid arthritis) can lead to the release of lysosomal enzymes into the extracellular space. This destroys the surrounding tissues. An imbalance between proteases and antiproteases can lead to destruction of lung tissue in emphysema caused by tobacco smoking.
Other diseases include muscular dystrophy, skin degeneration, respiratory and gastrointestinal diseases, malignant tumors.
Non-enzymatic proteolysis
The protein backbone is very stable in water at neutral pH and room temperature, although the rate of hydrolysis of different peptide bonds may vary. The half-life of peptide bond decay ranges from 7 to 350 years.
Strong mineral acids can easily hydrolyze peptide bonds in proteins. The standard way to hydrolyze a protein is to heat it to 105°C or soak it in hydrochloric acid for 24 hours.
Method of determination
There are several methods for determining proteolytic activity. For example, hydrolysis of casein, hemoglobin or azocasein. The first method is not expensive, but casein is difficult to dissolve. The hemoglobin hydrolysis method is more expensive. When using it, the substrate must be denatured. The third method avoids this, but it's also not cheap. The fastest, not expensive method is the use of a milk substrate. It includes less equipment and can be used in training courses. All it takes is skimmed milk and a water bath.
Experimental procedure
Two milliliters of a buffer solution (sodium acetate pH 5.0 containing CaCI2) is added to 3 milliliters of skimmed milk. This mixture is kept at 30 °C in a water bath for 10 minutes. To see the milk coagulation process, a light source is used. It skips for as many seconds as it takes to coagulate a piece of milk the size of a pinhead. An adequate period for accuracy is between one and two minutes. Enzyme block is defined as the amount required to form the first coagulated fragment in one minute under the chosen experimental conditions.
Proteases as antiviral agents
Currently, there are a number of approved drugs with proteolytic activity for use in the treatment of viral infections. Most are used primarily to treat herpesvirus, human immunodeficiency virus, respiratory syncytial infections, and influenza A virus infections. These are nucleoside analogs that act by inhibiting viral DNA synthesis.
Research over the past decade has shown that proteases are an absolute requirement in the life cycle of many viruses. Influence occurs eitherby cleavage of high molecular weight precursor proteins to obtain functional products, or by catalysis of structural proteins necessary for the assembly and morphogenesis of viral particles.
To date, four protease inhibitors have been approved:
- "Saquinavir" (Invirase, Ro 31-8959).
- Indinavir (Crixivan, MK-639).
- "Ritonavir" (Norvir, AVT-538).
- "Nelfinavir" (Viracept, AG1343).
Other drugs
Picornavirus proteases constitute one of the largest families of medically important human pathogens. Enteroviruses are associated with various clinical syndromes, including upper respiratory tract disease, aseptic meningitis, encephalitis, myocarditis, hand, foot, and mouth diseases. In this case, proteases will help. Expectorants with proteolytic activity:
- "Trypsin".
- "Ribonuclease".
- "Chimozin"
Another potential antirhinovirus drug is Pleconaril.
