Proteases (proteinases, peptidases and proteolytic enzymes) play a very important role in human life. To date, more than 500 of these enzymes are 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 - the use of serine alcohol.
- Cysteine ββ- the use of cysteine ββthiol.
- Threonine - a turnover of threonine secondary alcohol.
- Aspartic - using aspartic carboxylic acid.
- Glutamine - with the use of carboxylic acid glutamate.
- Metalloprotease is the turnover of a metal, usually zinc.
- LiAZases of an asparagine peptide - asparagine is used in order to carry out an exclusion reaction (without requiring water).
Proteases were first assembled into 84 families, according to their proteolytic activity, and classified under 4 catalytic types:
- serine;
- cysteine;
- aspartic;
- metal.
Value
General proteolytic activity plays an important role in many body processes. This is fertilization, digestion, growth, maturation, aging and even death. Proteases regulate many physiological processes by controlling the activation of synthesis and the deterioration of protein foods. They play an important role in the reproduction and spread of viruses, bacteria and parasites, therefore, they are responsible for the effective transmission of diseases caused by pathogenic substances. These enzymes enable tumor cells to divide, fill the intercellular space and blood vessels of the lungs, and also spread in 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 so that the protease is activated only in the right place and context. Incorrect proteolytic activity can be very damaging to the body.
- Protein degradation. May occur intracellularly or extracellularly. It performs a number of functions: removes damaged and abnormal proteins; prevents their accumulation; serves to regulate cellular processes by removing enzymes.
- Digestion. Proteins derived from food are split into peptide chains due to pepsin, trypsin, chymotrypsin and elastase. To prevent inappropriate or premature activation of digestive enzymes (this can cause pancreatitis), they act as an inactive zymogen.
Enzymes
Proteolytic enzymes are present 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 protein cleavage is catalyzed. The two main groups are egzopeptides and endopeptidases. Inside the body, protein materials are initially attacked by pepsin. When protein is transmitted to the small intestine, it is partially digested by the stomach. Here it is exposed to proteolytic enzymes secreted by the pancreas. Pancreatic enzymes are then activated in the intestine, turning 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 find out their role within the pathogenesis of infectious diseases. The main attention was paid to the examination of lactic acid bacteria from various yoghurts and fermented milk. They are widespread in nature. These are lactobacilli, lactococci, bifidobacteria, streptococci, enterococci and sporolactobacteria. They are divided by species, subspecies, variants and strains.
Proteolytic activity is a very important characteristic of lactic acid bacteria. Bacterial proteases are enzymes that catalyze hydrolyspeptide 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
The 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. Inactive pepsinogen proenzyme is synthesized inside 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 stimulator of the parietal cells. 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
Abnormal proteolytic activity is associated with many diseases. In pancreatitis, leakage of proteases and their premature activation in the pancreas leads to self-ignition of the pancreas. People with diabetes can have increased lysosome activity, the degradation of some proteins can increase significantly. Chronic inflammatory diseases (rheumatoid arthritis) can lead to the release of lysosomal enzymes into the intercellular space. This destroys the surrounding tissue. An imbalance between proteases and antiproteases can lead to destruction of lung tissue with emphysema caused by tobacco smoking.
Other diseases include muscular dystrophy, skin degeneration, respiratory and gastrointestinal diseases, and malignant tumors.
Nonenzymatic proteolysis
The protein base 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 the binding of the peptide ranges from 7 to 350 years.
Strong mineral acids can easily hydrolyze peptide bonds in a protein. The standard way to hydrolyze a protein is to heat it to 105 Β° C or hold it for 24 hours in hydrochloric acid.
Determination method
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 method of hydrolysis of hemoglobin is more expensive. When using it, the substrate must be denatured. The third method avoids this, but it is 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 that is required is skim milk and a water bath.
Experimental procedure
Two milliliters of buffer solution (sodium acetate with pH 5.0 containing CaC I2 ) is added to 3 milliliters of skim milk. This mixture is aged at 30 Β° C in a water bath for 10 minutes. A light source is used to see the clotting process of milk. It is skipped for as many seconds as necessary to coagulate a milk particle the size of a pin head. An adequate period for accuracy is between one and two minutes. An enzyme block is defined as its amount, which is necessary for the formation of 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 of them are mainly used to treat herpes virus infections, human immunodeficiency virus, respiratory syncytial infections and influenza A. These are nucleoside analogues 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. The effect occurs either by splitting high molecular weight precursor proteins to produce 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, -538).
- Nelfinavir (Virasept, AG1343).
Other drugs
Picornavirus proteases are one of the largest families of medically important human pathogens. Enteroviruses are associated with various clinical syndromes, including diseases of the upper respiratory tract, aseptic meningitis, encephalitis, myocarditis, diseases of the hands, feet and mouth. In this case, proteases will help. Expectorants with proteolytic activity:
- Trypsin.
- "Ribonuclease".
- Khimozin
Another potential antirinovirus drug is Plekonaril.