BIOCHEMISTRY - L. Stryer - 1984

VOLUME 1

PART I. CONFORMATION AND DYNAMICS

CHAPTER 6. INTRODUCTION TO ENZYMOLOGY

6.8. Some Properties of Active Sites

The Active Site of an enzyme is the region that binds substrates (and the prosthetic group, if present) and contains amino acid residues directly involved in the formation or Cleavage of chemical bonds. Such residues are referred to as catalytic groups. Despite the enormous diversity of enzyme structures, their specificities, and catalytic mechanisms, several General Principles regarding The properties of active sites can be established.

1. The active site occupies a relatively small fraction of the total volume of the enzyme. The majority of amino acid residues in an enzyme molecule do not make contact with the substrate. It remains an enigma why Enzymes are so large. Nearly all enzymes contain more than 100 amino acid residues, have a molecular mass exceeding 10 kDa, and a diameter greater than 25 Å.

2. The active site is a three-dimensional entity. In other words, it is not a point, a line, or even a plane, but rather a complex three-dimensional Structure formed by groups originating from different PARTS OF THE linear Amino Acid Sequence. Indeed, as we have already seen in the Examples of Hemoglobin and Myoglobin, interactions between amino acid residues situated far apart in the primary sequence are frequently stronger than interactions between adjacent residues. In Lysozyme—an enzyme we will examine in detail in the next chapter—the key residues of the active site are located at positions 35, 52, 62, 63, and 101 within the 129-amino-acid linear sequence.

3. Substrates are bound to enzymes relatively weakly. Equilibrium constants for ES complexes typically range from 10^-2 to 10^-8 M, corresponding to standard free energies of interaction between — 3 and — 12 kcal/mol. For comparison, the strength of covalent bonds ranges from — 50 to — 110 kcal/mol.

4. The active site takes the form of a narrow cleft or crevice. In all enzymes of known structure, substrate binding occurs within such a cleft or crevice, which is largely excluded from Water except when water acts as a reactant. Several polar amino acid residues essential for binding and catalysis are located within this pocket. The generally nonpolar Nature of the surrounding region promotes substrate binding. Furthermore, the cleft-like geometry of the active site creates a microenvironment in which specific polar residues acquire unique properties vital for catalysis.

5. Binding Specificity depends on the precise spatial arrangement of atoms within the active site. A substrate can enter the active site only if its shape is complementary to it. In 1890, Emil Fischer introduced the lock-and-key model (Fig. 6.10), which proved to be fundamentally sound and an exceptionally fruitful concept for understanding the stereospecificity of catalysis. However, as recent studies show, the active sites of certain enzymes are not rigid structures; their conformation is modified upon substrate binding. In these enzymes, the active site becomes complementary to the substrate in accordance with the induced-fit model. Substrate binding induces a conformational change in the enzyme. The active site becomes complementary to the substrate's shape only after the substrate is bound. This dynamic recognition process is termed induced fit (Fig. 6.11). Moreover, some enzymes preferentially bind substrates in a strained ("distorted") conformation that resembles the Transition State.

Class="center">Fig. 6.10. Interaction of substrates with enzymes According to the lock-and-key model. The active site of the enzyme is inherently complementary in shape to the substrate

Fig. 6.11. Substrate interaction