BIOCHEMISTRY: A TEXTBOOK FOR UNIVERSITIES - E. S. Severin - 2004

SECTION 2. ENZYMOLOGY

Chemical processes form the foundation of life in any Organism. Virtually all reactions in living systems proceed with the participation of natural biocatalysts known as Enzymes. Among the vast array of energetically feasible reactions, enzymes selectively channel reactants, referred to as substrates, along physiologically useful pathways. Thus, enzymes govern all metabolic processes within the organism.

In Russian-language scientific literature, both terms—"ferment" and "enzyme"—have become firmly established, though preference is generally given to "ferment," even though the science itself is called enzymology. The word "ferment" originates from the Latin fermentum (leaven), while "enzyme" derives from the Greek en (in, within) and zyme (Yeast). This terminology emerged historically during The Study of the enzymatic processes of Alcoholic Fermentation.

Enzymology emerged as a distinct science in the early 19th century, and its active development continues to this day. Its objectives include determining the specific role of individual enzymes in accelerating Chemical Reactions within the organism, isolating and purifying enzymes, elucidating their structures, investigating Mechanisms of action, and studying kinetic characteristics and The regulation of activity in vivo.

For practical medicine, Structure/19.html">The Importance of enzymology lies in providing pharmacologists with a tool to purposefully alter cellular METABOLISM by targeting enzyme activity with specific chemical agents. A vast number of pharmaceutical drugs function as Enzyme Inhibitors. Another equally vital application of enzymology in clinical practice is The Use of enzyme activity assays in biological fluids for disease Diagnosis. Furthermore, isolated and purified Enzymes can be employed as therapeutic agents.

I. General Characteristics of Enzymes as Biological Catalysts

As established as early as 1922, enzymes are Proteins. Their role is unique: they increase The rate of chemical reactions without being consumed in the process. In 1926, urease—an enzyme catalyzing The breakdown of urea into ammonia and carbon dioxide—was first purified and isolated in the form of protein crystals. To date, hundreds of different enzymes have been obtained in crystalline form, their Amino acid sequences have been decoded, and their roles in metabolic transformations are actively being investigated.

Non-protein compounds can also act as biocatalysts. For instance, certain types of RNA catalyze the Hydrolysis of phosphodiester bonds in Nucleic Acids. Such catalytically active RNA molecules are termed ribozymes, although their significance in the chemical transformation of compounds is far smaller than that of protein enzymes.

Since enzymes are protein molecules, they possess all the properties characteristic of proteins. At the same time, they exhibit structural features that define them as catalysts. Let us examine the Fundamental properties of enzymes as biological catalysts.

A. Specificity

The biological function of an enzyme, like that of any protein, is determined by the presence of an Active Site in its structure. A Ligand that interacts with the enzyme's active site is called a substrate. The active site contains amino acid residues whose functional groups ensure substrate binding, alongside residues whose functional groups execute the chemical transformation of the substrate. Conventionally, these are designated as the binding site and the catalytic site, though it should be remembered that these regions do not always have a rigid spatial Separation and may occasionally overlap (Fig. 2-1).

Class="center">Fig. 2-1. STRUCTURE OF THE enzyme active site. A – binding of the substrate to the enzyme at the active site; B – arrangement of The amino acid residues forming the active site within the Introduction/19.html">Primary Structure of the protein; C – conventional division of the enzyme active site into a binding site and a catalytic site. The binding site is formed by amino acid side chains whose functional groups ensure substrate binding. The catalytic site is formed by amino acid residues whose functional groups facilitate the chemical Conversion of the substrate.

Within the binding site, the substrate interacts (binds) with the enzyme via non-covalent bonds, forming an enzyme-substrate complex. In the catalytic site, the substrate undergoes a chemical transformation into a product, which is subsequently released from the enzyme's active site. Schematically, the catalysis process can be represented by the following equation:

Е + S <-> ES <-> ЕР <-> Е + Р,

where E is the enzyme, S is the substrate, and P is the product. These designations are universally accepted and originate from the English words enzyme, substrate, and product.

Specificity is the most crucial property of enzymes, determining the Biological Significance of these molecules. One distinguishes between substrate and catalytic specificities of an enzyme, both of which are governed by The structure of the active site (Fig. 2-2).

Fig. 2-2. Functional Significance of individual Regions of the enzyme active site.

1. Substrate Specificity

Substrate specificity refers to the ability of each enzyme to interact with only one or a few specific substrates. The following types are distinguished:

✵ absolute substrate specificity;

✵ group substrate specificity;

✵ stereospecificity.

Absolute Substrate Specificity

The Active Site of enzymes with absolute substrate specificity is complementary to only a single substrate. It should be noted that such enzymes are rare in living organisms.

An example of an enzyme with absolute substrate specificity is arginase, which catalyzes the Cleavage of Arginine into urea and Ornithine:

Another example of an enzyme with absolute substrate specificity is urease, which catalyzes the hydrolysis of urea into carbon dioxide and ammonia.

Most enzymes catalyze similar types of reactions involving a small group of structurally related substrates.

For instance, the enzyme pancreatic lipase catalyzes the Digestion of dietary fats in the human duodenum, facilitating The conversion of any fat molecule (triacylglycerol) into one molecule of monoacylglycerol and two molecules of Higher Fatty acids. Pancreatic lipase hydrolyzes the ester bond at the α-carbon atoms of glycerol, regardless of the specific fatty acids composing the fat molecule (see scheme).

Scheme

Most Proteolytic Enzymes involved in Protein Hydrolysis exhibit group substrate specificity, cleaving peptide bonds formed by various Amino Acids.

Stereospecificity

When a substrate has multiple stereoisomers, an enzyme displays absolute specificity toward only one of them. In The Human Body, Enzyme Specificity is observed for the following stereoisomers.

Stereospecificity for D-sugars. The majority of Monosaccharides AND THEIR metabolic products in humans and other mammals belong to the D-stereoisomers. The enzymes responsible for their metabolism are specific to D-sugars rather than L-sugars.

Stereospecificity for L-amino acids. Human proteins are composed of L-amino acids. Most enzymes involved in amino acid transformations exhibit stereospecificity for L-amino acids.

Stereospecificity for cis-trans isomers. The enzyme fumarase acts exclusively on fumarate. Maleate (the cis-isomer of fumarate) does not serve as a substrate for fumarase.

The only exceptions are epimerases (racemases), which catalyze the interconversion of optical isomers.

Stereospecificity for α- and β-glycosidic bonds. The enzyme amylase acts solely on α-glycosidic bonds, enabling the Hydrolysis of Starch and Glycogen (glucose polymers), in which glucose residues are linked by α-glycosidic bonds. Cellulose is also a glucose polymer; however, its glucose residues are linked by β-glycosidic bonds. Because humans lack enzymes specific to β-glycosidic bonds, cellulose is not digested in the human intestine and cannot serve as a source of glucose.

2. Catalytic Specificity

An enzyme catalyzes the conversion of a bound substrate via one specific pathway out of several possible routes. This property is determined by the structure of the catalytic region within the enzyme's active site and is referred to as catalytic specificity, or reaction pathway specificity. For example, in human Liver Cells, glucose-6-phosphate serves as a substrate for four different enzymes: phosphoglucomutase, glucose-6-phosphatase, phosphoglucose isomerase, and glucose-6-phosphate dehydrogenase. However, due to structural differences in the catalytic sites of these enzymes, the compound undergoes distinct transformations yielding four different products (see the scheme below).

B. Catalytic Efficiency

Most enzyme-catalyzed reactions are highly efficient, proceeding 10⁸ to 10¹⁴ times faster than uncatalyzed reactions. Each enzyme molecule is capable of transforming anywhere from 100 to 1,000 substrate molecules into product per second.

The number of substrate molecules converted into product by a single enzyme molecule per second is known as the turnover number, or catalytic center activity.

V. Enzyme liability

The catalytic efficiency of an enzyme, like that of any protein molecule, depends on its conformation, particularly the conformation of its active site.

Enzymes are characterized by conformational lability, which is the capacity for subtle alterations in native conformation resulting from the disruption of weak bonds. Consequently, exposure to Denaturing Agents capable of altering the enzyme molecule's conformation leads to A change in the active site's conformation and a reduced ability to bind the substrate. As a result, the catalytic efficiency of the enzyme decreases.

G. Regulatory Properties of Enzymes

Enzyme activity within The Cell depends on the concentration of substrate and product molecules, as well as the availability of Cofactors and Coenzymes. Cellular enzymatic processes are typically strictly ordered: the product of one enzymatic reaction serves as the substrate for another, thus forming metabolic pathways. Within most metabolic pathways, specific key or regulatory enzymes can be distinguished, whose activity adapts to the cell's demand for the pathway's end product. Regulatory enzymes are generally located at the beginning of a metabolic pathway and/or at branch points. They catalyze either the slowest (rate-limiting) reactions or irreversible reactions. A detailed Discussion of how Metabolic control is achieved through the Regulation of enzyme Activity is provided in subsection VII.