Chapter Introduction

5: Protein Function

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  • 5.1 Protein-Ligand Interactions

  • 5.2 Enzymes: The Reaction Catalysts of Biological Systems

  • 5.3 Motor Proteins

  • 5.4 The Regulation of Protein Function

MOMENT OF DISCOVERY

Smita Patel

My story shows the insights one can achieve by using the right tools. During my postdoctoral years with Ken Johnson, I studied the DNA polymerase of bacteriophage T7. Charles Richardson demonstrated that replication of the T7 chromosome requires a DNA helicase, encoded by the T7 genome as gene 4. Collaborating with Bill Studier (who had recently constructed the first of a set of excellent cloning vehicles called pET vectors), I cloned and expressed gene 4. With this clone, I was able to purify the gene 4 protein product. I took this with me when I started my first faculty position at Ohio State University in 1992.

In the early days of getting my lab set up, I ran the lab with a small army of undergraduates. These students carried out a range of assays on the T7 DNA helicase. The enzyme bound stoichiometrically to DNA oligonucleotides that were 10 to 20 nucleotides in length, and hydrolyzed TTP and ATP. However, we were confused that the binding stoichiometry was consistently six to seven monomers of T7 helicase to each DNA molecule. We initially thought we had quite a lot of inactive enzymes.

Helicases were fairly new enzymes in those days. The DNA helicases getting the most attention were monomers or dimers in their active form. The RNA helicase Rho was known to be a hexamer, as was the DnaB helicase, but this did not seem to be a pattern. Then, my first graduate student, Manju Hingorani, decided to look at the protein directly by electron microscopy. This finally answered our questions. I remember my excitement as we looked at the beautiful ring structures in the electron micrographs. We could count up to five subunits. Later, collaborating with Ed Egelman, higher-resolution micrographs helped to establish that T7 helicase was a hexamer and the central channel was binding the DNA. We realized that rings are general in nature and widely used as replicative helicases that travel through thousands of base pairs of DNA without falling off. We published the results in a series of papers from 1993 to 1995. Since then, I have been endlessly captivated by these fascinating ring-shaped motor proteins.

—Smita Patel, on her early work with the T7 gene 4–encoded DNA helicase

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Biological information—in the form of the genome of every organism and virus—is the focus of molecular biology, and of this textbook. As is true for all cellular functions, the packaging, function, and metabolism of this genomic information involve a wide range of proteins and RNA molecules. These macromolecules can be broadly divided into three classes, depending on function: reversible binding, catalysis, or motor activities. In the first three sections of this chapter, we review these macromolecular functions in succession, focusing on their roles in DNA and RNA metabolism. First, some proteins and RNAs simply bind reversibly to nucleic acids; this binding often has a structural or regulatory function. Second, another large class of proteins (and some RNAs) act as biological catalysts, accelerating the reactions needed to sustain and propagate living systems. These are the enzymes, as critical to life as are the information-containing DNA and RNA genomes. And third, motor proteins do the work of moving cellular molecules from one location to another, of separating molecules, and of bringing molecules together.

The great majority of macromolecules that carry out these three functions are proteins, although several RNA enzymes are known and are increasingly well understood. The functions of proteins are particularly important to the topics of every chapter in this book, and an introduction to protein function now becomes our focus. The various functions of RNAs are described in Chapters 15 and 16, although the general principles described here apply to RNA molecules as well as to proteins. In this chapter, after exploring each of the three major functions of proteins, we conclude with a discussion of protein regulation.