Molecule in the cell are in constant motion, and these random thermal motions cause them to come into contact with one another on a frequent basis. The poorly matched surfaces of two molecules that collide produce minimal noncovalent bonds, and the two molecules almost instantly split following the collision. On the other hand, the connection may continue for a very long time when several noncovalent bonds occur between two molecules that have been in collision with one another. Strong interactions take place in cells when a biological process requires that molecules remain linked for an extended period of time. For example, when a collection of RNA and protein molecules come together to form a subcellular structure like a ribosome, these molecules must remain linked for the duration of the process.

The degree to which any two molecules are able to bind to one another can be quantified. Imagine a fluid containing a population of identical antibody molecules, all of which at some point get suddenly exposed to a population of ligands that are moving through the fluid in the area around them. The binding site on an antibody will commonly come into contact with one of the ligand molecules, which will result in the formation of an antibody-ligand combination. Because of this, the number of antibody-ligand complexes in the population will increase, but this growth will not continue indefinitely. A second phase, in which individual complexes disintegrate as a result of thermally induced motion, will over time become of greater relevance. This process will take place over time. Any population of antibody molecules and ligands will, with time, move closer and closer to a stable state, also known as equilibrium. This state is characterized by a precise balance between the number of "binding" (association) and "unbinding" (dissociation) events.

By computing the equilibrium constant from the concentrations of the ligand, antibody, and antibody-ligand complex at equilibrium, we may derive a straightforward method for determining how strongly two molecules are bound to one another (K). According to the equilibrium constant for a reaction in which two molecules (A and B) bind to one another to create a complex, when the concentration of a ligand (measured in moles/liter) reaches a value equal to 1/K, half of the binding sites will be occupied by the ligand. This is the case when the concentration of the ligand is high enough (AB). This equilibrium constant, which directly quantifies the difference in free energy between the bound and free states, grows larger as the binding strength of the system grows stronger. Even a slight change in the way noncovalent connections are made can result in significant alterations to the binding interactions between molecules.

Using the example of an antibody binding to its ligand, it was explained how the strength of the binding might affect the equilibrium state, but the same concepts are applicable to any molecule and its ligand. As we will see in the following section, many proteins are enzymes that, once they have bound to their respective ligands, accelerate the production of covalent bonds in these molecules or catalyze the breaking of covalent bonds already present.

The majority of tasks that proteins are designed to perform can be accomplished by simply attaching themselves to another molecule. By way of illustration, an actin molecule can initiate the formation of a filament by merely combining its forces with those of additional actin molecules. On the other hand, the binding of ligands is merely the initial, but critically important stage in the function of some proteins. This is the case with enzymes, which make up a massive and extremely important category of proteins. Enzymes are remarkable molecules that are responsible for the chemical events that build and dissolve covalent connections in cells. Enzymes are found in all living organisms. They then connect themselves to one or more ligands, which are also referred to as substrates, and turn those ligands into one or more products that have undergone chemical modification. Enzymes play the role of catalysts, enabling cells to make or break covalent connections in a manner that is under their direct control. They have the ability to accelerate reactions by a factor of a million or more without affecting the reactions themselves in any way. Catalyzing organized sequences of chemical reactions is what makes life possible and is done by enzymes, which are also responsible for the production and maintenance of cells.

The chemical reactions that enzymes catalyze allow for their classification into a variety of different functional types. Because of their high specialization, the enzymes that belong to this class can each only catalyze a single type of chemical reaction. Therefore, the enzyme that causes blood to clot, known as thrombin, can only break one type of blood protein, which occurs between a particular arginine and its neighboring glycine, and nowhere else. On the other hand, it can add a phosphate group to D-glucose while ignoring its optical isomer, L-glucose, but it cannot break any other type of blood protein.

The attachment of a substrate molecule to a protein that is responsible for catalyzing a chemical reaction (an enzyme) is a prerequisite that must be met. The simplest route for a reaction to take is denoted by the notation E + S ES EP E + P, where E represents the enzyme, S represents the substrate, and P represents the product. There is a limit to the amount of substrate that a single enzyme molecule can process in a given amount of time. In spite of the fact that the pace at which product is created speeds up as the concentration of the substrate rises, this rate eventually achieves its maximum possible value. Because the enzyme molecule is already at its maximum capacity of substrate at that point, the rate of reaction, also known as Vmax, is only dependent on how rapidly the enzyme can break down the substrate molecule. Calculating the turnover number requires taking the maximal rate and dividing it by the concentration of the enzyme. Although turnover values ranging from one to ten thousand have been documented, the typical rate at which one enzyme molecule may process around one thousand substrate molecules in one second is one.

The concentration of substrate at which the reaction can proceed at a rate that is half of its highest possible rate (0.5 Vmax) is referred to as an enzyme's Km. This is another kinetic parameter that is commonly used to define an enzyme. A low Km value indicates that the enzyme binds to its substrate very strongly and that it reaches its maximum catalytic rate at low substrate concentrations, whereas a high Km value indicates that the enzyme binds to its substrate in a relatively weak manner.

The rates at which enzymes catalyze chemical processes are orders of magnitude higher than those at which any artificial catalyst can catalyze chemical reactions. There are a variety of reasons that contribute to this high level of productivity. When there are two molecules that need to react, the enzyme first drastically increases the local concentration of the two substrate molecules at the catalytic site and then retains them there in the correct orientation for the subsequent reaction. But what's even more important is that some of the energy used to bind the molecules is directly involved in the catalysis. Before the substrate molecules can proceed to create the ultimate products of the reaction, they must first transition through a variety of intermediate states, each of which has a different size, shape, and distribution of electrons. The activation energy of a reaction is the free energy that is required to reach the transition state, which is the intermediate stage that is the most unstable. The activation energy is an important factor in determining the pace at which a reaction occurs. The affinity of enzymes for the transition state of the substrate is substantially higher than their affinity for the substrate in its stable form.

Enzymes are the most efficient and selective of all the catalysts.

In order to understand the chemistry of cells and animals, as well as the synthesis of valuable substances on an industrial scale and the discovery of new drugs, it is essential to have a thorough comprehension of their complex operations. By carefully analyzing the rates of the chemical reactions that are catalyzed by a purified enzyme, and more specifically how these rates change with changes in conditions such as the concentrations of substrates, products, inhibitors, and regulatory ligands, biochemists are able to determine the precise mechanism of action that is utilized by each enzyme.