Enzyme

I

Introduction

Enzyme, a protein that catalyses a specific reaction. Living cells contain thousands of different enzymes, each of which catalyses (that is, accelerates without itself being changed) just one kind of reaction. In some of these reactions, small organic molecules such as amino acids, sugars, nucleotides, and lipids are broken down to provide energy for the cell. In other reactions, small molecules are built into complex macromolecules, such as proteins, DNA, RNA, and polysaccharides, or used to carry signals, or to control cell movements or gene expression. Enzyme-catalysed reactions are usually connected in series, so that the product of one reaction becomes the starting material, or substrate, for the next. These long linear reaction pathways are in turn linked to one another, forming a maze of interconnected reactions that enables the cell to survive, grow, and reproduce. Enzymes exhibit enormous catalytic power, in some cases increasing reaction rates by a factor of over 10¹⁴ (one hundred million million). Enzymes dictate the pattern of chemical changes in a cell and without them life as we know it would be impossible.

II

Discovery of Enzymes

Enzymatic processes such as alcoholic fermentation and the digestion of meat have been known since antiquity. Until the end of the 19th century, when the German chemist Eduard Buchner discovered that a cell-free extract of yeast can ferment glucose to alcohol and carbon dioxide, such phenomena were believed to dependent on the presence of living organisms. This seminal finding led to a search for the many biochemical breakdown products of glucose in yeast extracts. The agents causing these transformations came to be known as enzymes, from a Greek phrase meaning 'in ferment'. In 1926, the American biochemist James Sumner succeeded in isolating and crystallizing the enzyme urease, which converts urea to carbon dioxide and ammonia. Four years later pepsin and trypsin, two enzymes from the digestive system that break down proteins, were isolated and crystallized by the American biochemist John Northrop. Enzymes were found to be proteins and Northrop proved that the protein was actually the enzyme and not simply a carrier for another compound.

Research in enzyme chemistry in recent years has shed new light on some of the most basic functions of life. Ribonuclease, a simple enzyme that degrades molecules of RNA was discovered in 1938 by the American bacteriologist René Dubos and isolated in 1946 by the American chemist Moses Kunitz. The molecule was synthesized by American researchers in 1969 in a synthesis that involved hooking together 124 amino acids in a very specific sequence to form protein molecules. Such syntheses led to the probability of identifying those areas of the molecule that carry out its chemical functions, and opened up the possibility of creating specialized enzymes with properties not possessed by the natural substances. This potential has been greatly expanded in recent years by genetic engineering techniques that make it possible to produce enzymes in great quantities (see Biochemistry).

III

How Enzymes Work

The chemical reactions that a cell performs would normally occur only under extreme conditions such as at high temperatures and in non-aqueous solvents. The reacting molecules have to overcome an energy barrier before they can react, which is difficult to do in dilute aqueous solution and at body temperature. Enzymes achieve this feat by binding to the reacting molecules and by holding them close together in the precise orientation needed for the desired reaction to occur.

Enzymes from mammalian sources usually operate optimally at body temperature and are easily destroyed (or 'denatured') by heating at temperatures above about 70°C. However, there are enzymes—such as those from thermophilic bacteria from deep-sea thermal vents (see Hydrothermal Vent)—that are stable even at temperatures close to boiling water (and are widely used by biochemists because of their great stability).

To see how enzymes work, consider the example of the enzyme hexokinase, which catalyses the transfer of a phosphate group from ATP (adenosine 5'-triphosphate) to the sugar glucose. ATP is a small molecule that carries chemical energy in cells. The transfer of one of its phosphate groups to a sugar such as glucose is the first stage in the metabolic breakdown of the sugar molecule. Although the reaction is chemically favourable, it will not occur under the conditions existing in a living cell unless there is a specific enzyme present to speed it up. The protein hexokinase binds ATP and glucose in such a way that the energy barrier to the transfer of a phosphate group is dramatically reduced.

The hexokinase molecule has evolved a pocket or groove on the protein's surface, known as its active site, into which glucose and ATP fit snugly. Glucose and ATP bind tightly to the enzyme and with exactly the correct relative positions for the transfer of a phosphate group to occur. Moreover, other amino acid side chains in the binding site interact transiently with the two reacting molecules so as to help the reaction. Once the reaction is completed, then the enzyme releases the products, which are allowed to diffuse away. The enzyme is then ready to catalyse another reaction of the same kind.

IV

Allosteric Enzymes

If the structure of the enzyme hexokinase is examined in atomic detail it is found that the protein molecule has two compact regions, or domains, joined by a flexible linker. These two domains are capable of independent movement, and can open and shut rather like a pair of jaws. When glucose and ATP bind to the enzyme, the two domains close around them, thereby ensuring a very tight fit. Once the reaction is complete, the jaws open again to release the products phosphoglucose and ADP.

Small movements such as these, termed conformational changes, contribute to the catalytic process of many enzymes. They are also an important factor in the regulation of enzyme activity. It is vitally important to a cell that its many reactions take place only when required. To this end, many enzymes can be switched on and off according to conditions in the cytoplasm. In one form of control known as feedback inhibition (see Homeostasis) the final product of a long series of enzyme reactions binds to and inhibits an enzyme at the start of the pathway. By limiting entry into the pathway in this way, feedback inhibition prevents a wasteful build-up of intermediates. Enzymes are also subjected to positive regulation and can be activated by the binding of a small molecule or by a chemical modification such as the addition of phosphate group.

The substances that inhibit or activate an enzyme are often unrelated chemically to its substrates. They bind to locations on the enzyme surface other than its active site and influence the catalytic mechanism by causing a conformational change in the enzyme. For example, the binding of a small molecule might cause a conformational change that opens up an active site that is otherwise closed and thereby switch an enzyme on. Enzymes that are regulated in this fashion have two distinct binding sites on their surface and are referred to as being allosteric meaning 'other shape' for this reason. Allosteric enzymes are typically built from more than one protein chain and in many cases have a symmetrical structure. Conformational changes in one sub-unit may then influence the changes in adjacent sub-units of similar structure leading to a sharper and more switch-like response.