Introduction

D.A.S. Grahame1; B.C. Bryksa1; R.Y. Yada2    1 University of Guelph, Guelph, ON, Canada
2 University of British Columbia, Vancouver, BC, Canada

1.1 Introduction

The myriad of proteins and enzymes encoded for by the genetic code represents a portion of the realized potential of a genetic blueprint. Within this genetic blueprint lies the code for enzymes responsible for vital processes such as regulating blood pressure, aiding digestion, and supporting growth and development. As with many things, having an understanding and knowledge of the foundation on which current studies and techniques rely helps to ensure an appreciation for, and an ability to, fully comprehend the theory contained within. In that regard we will briefly introduce the history of enzymes and enzymology in an attempt to give a basic background on which to build in the chapters to come.

1.1.1 Historical perspective on enzymes

One of the first recorded uses of enzymes comes from Homer’s Iliad in 850 BC, wherein milk was transported in the stomach of a young goat to produce cheese. Alternatively, in ancient China, enzymes such as amylase and various proteases were used as digestive aids (Adrie and Straathof, 2000; Zhu and Tramper, 2013). Like China, Japan has a long history of enzyme use, most often exemplified in the form of koji, a fungi-fermented wheat or rice product that produces the enzyme amylase. Koji is important because it is a required component for the production of rice wine, soy sauce, soybean products and other distilled alcoholic spirits (Zhu and Tramper, 2013). However, it was not until the late 1700s and early 1800s, when Spallanzani reported on the action of proteases and Lavoisier and Gay-Lussac published their works on the chemistry of fermentation, that significant work began on processes dominated by enzymes (Buchholz and Collins, 2013). In fact, it is generally regarded that the term enzyme, as well as a fundamental understanding of the basic unit of an enzyme, was not achieved until the mid to late 1800s.

To understand how the work of individuals such as Spallanzani and Lavoisier played a role in the development of enzymatic theory, one must look at the work done by Pasteur and Liebig on the fermentation of sugar to alcohol in the 1850s. Pasteur postulated that the fermentation of sugar to alcohol occurs due to the action of “living yeast.” Liebig, on the other hand, believed that the fermentation of sugar to alcohol was a non-living process and purely chemical in nature. Although Pasteur and Liebig’s conclusions were contradictory, both are dependent on the work of Schwann and Danilewski in 1837 (involving pepsin and trypsin), which details living cells as fermentation units (Buchholz and Collins, 2013). As for Pasteur and Liebig, the findings of Schwann and Danilewski had built on the work of Kirchhoff in 1814 that describes the ability of wheat to convert starch to sugar. The work of Pasteur and Liebig caused a ferocious debate regarding whether fermentation should be considered a living or a non-living process. This debate eventually led to the involvement of numerous scientists such as Kuhne who is credited with being the first individual to propose the term enzyme (in 1876) to describe the unorganized fermentation seen in the work of Pasteur and Liebig. Unfortunately for Pasteur and Liebig, the debate was settled only after both of their deaths; in 1897, the Buchner brothers discovered and demonstrated the ability of sugar to be converted to ethanol using yeast extracts instead of whole yeast cells. This breakthrough showed that the conversion or fermentation of sugar was the action of non-living components of cells. Thus the answer to the question of how the work of Spallanzani and Lavoisier, among others, eventually led to the generation of enzymatic theory is that other researchers built on their work through generations of scientific inquiry and vigorous debate.

The study of how and why enzymes are capable of conducting their modes of action accelerated in the late 1800s. In fact it was in a study of the enzyme invertase by O’Sullivan and Thompson (1880–1890) in which the first semblance of enzymatic kinetics began to appear. Unfortunately for O’Sullivan and Thompson, their work was incomplete, and it wasn’t until after the work of Henri (1889) and Fischer (1894) that an initial understanding of both the general rate law for enzyme kinetics, and a general hypothesis about how a substrate and enzyme bind, were formed. The work done by Fischer and Henri ended up being vital to the work of Brown (1902), who defined the enzyme–substrate complex in terms of enzyme kinetics. At this point a general theory involving the rates of enzymes, the binding of substrate, and the ability to define the enzyme–substrate complex in the kinetic reaction was available and necessary for Michaelis and Menten to propose their classic theory of enzyme kinetics, which they did in 1913. A number of enzymes were subsequently identified and characterized, leading to the first crystallization of a pure enzyme by Summer just 29 years after the Buchner brothers’ breakthrough.

With a theory of enzyme kinetics and the ability to crystallize protein, another rapid explosion of research involving enzymatic processes followed. In fact, rapid advances in the study and analysis of protein structures in the 1960s through the 1990s not only provided further insight into the theory proposed by Michaelis and Menten, but also gave rise to additional theories involving the quantum nature of enzymes as well as to three-dimensional modeling of enzymes in silico. The above text briefly covers the history of enzymology and hopefully demonstrates the massive amount of time and work on which the knowledge detailed in the following chapters is based.

1.1.2 Some of the basics

As was alluded to in our brief introduction to the history of enzymes, their uses in the generation of food and beverages, their ability to generate components of a food system, and their involvement in the processing and digestion of food, encompasses a diverse set of enzymes. Before we begin a brief introduction to enzymology as it relates to the food industry, we will first discuss how to classify and name enzymes.

1.1.2.1 Classification of enzymes

The rampant study of enzymes in the twentieth century led to a large number of novel enzymes being discovered. An unintended problem arose with the rapid discovery of these novel enzymes: The lack of a formal naming system soon caused individual enzymes to be known by several different names (Cornish-Bowden, 2014). As a result, the 1955 General Assembly of the International Union of Biochemistry set up an international commission to examine enzymes, and the commission subsequently created a classification system for enzymes that is still in use today. The classification system proposed by the commission recommends that enzymes be classified using three general principles (Cornish-Bowden, 2014; Enzyme Nomenclature, 2014):

• Names purporting to be names of enzymes, especially those ending in -ase, should be used only for single enzymes.

• Enzymes are principally classified and named according to the reaction they catalyze.

• Enzymes are divided into groups based on the substrate acted upon.

The scheme proposed contains four numbers separated by periods (e.g., 1.1.1.1), and each number in the scheme defines a particular characteristic. The initial number is the most important and separates an enzyme into one of six major types (Cornish-Bowden, 2014):

Class 1—Oxidoreductases: Catalyze reactions in which a substrate donates one or more electrons to an electron acceptor, becoming oxidized in the process.

Class 2—Transferases: Catalyze reactions in which a chemical group is transferred from a donor substrate to an acceptor substrate.

Class 3—Hydrolases: Catalyze reactions in which a bond in a substrate is hydrolyzed to produce two fragments.

Class 4—Lyases: Catalyze non-hydrolytic reactions in which a chemical group is removed from a substrate, leaving a double bond.

Class 5—Isomerases: Catalyze one-substrate, one-product reactions that can be regarded as isomerization reactions.

Class 6—Ligases: Catalyze the joining together of two or more molecules coupled to hydrolysis of ATP or an analogous molecule. These enzymes are also sometimes called synthetases.

With an understanding of how we classify and appropriately name enzymes, we will now introduce the major enzymes used in the food industry.

1.1.2.2 Common food enzymes

Traditionally, the most commonly used enzymes in the food industry are those applied during the production of cheeses and beverages such as beer. However, with the explosion of research about various enzymes, it has become apparent that the number of enzymes involved in the production of different food products is enormous.

Alpha-amylase: Hydrolyzes the alpha bonds of alpha-linked polysaccharides, producing glucose and maltose. It has multiple uses, including in baking and brewing, but it may be best known for producing corn syrup.

Beta-glucanase: Breaks down glucans in malt and other materials. Beta-glucanase is a key component in extract...