Protein Structure and Function PDF

Chapter 3 Protein Structure and Function Computationally designed, hypothetical four-stranded fiber protein viewed down the fiber axis. Using advanced methods for protein design, the amino acid sequence of the staphylococcal nuclease protein was modified so that the individual, folded protein chain...

Chapter 3 Protein Structure and Function Computationally designed, hypothetical four-stranded fiber protein viewed down the fiber axis. Using advanced methods for protein design, the amino acid sequence of the staphylococcal nuclease protein was modified so that the individual, folded protein chains (shown in different colors) would stack into long strands. The design predicts that four long strands would assemble into a four- stranded fiber held together by the binding of hydrophobic helices located in the center of the fiber. [Data from H. Shen et al., 2018, Science 362:705–709.] Outline 3.1 Hierarchical Structure of Proteins 3.2 Protein Folding 3.3 Protein Binding and Enzyme Catalysis 3.4 Regulating Protein Function 3.5 Purifying, Detecting, and Characterizing Proteins 3.6 Proteomics Proteins, which are polymers of amino acids called polypeptides, fold into three- dimensional structures of many sizes and shapes. Their three-dimensional diversity principally reflects variations in their amino acid sequences and in the lengths of the polypeptides. In general, a polypeptide will fold into only one or a few closely related three-dimensional shapes — called conformations. A key concept in understanding how a protein works is that its function is often derived from its three-dimensional structure, and its three-dimensional structure is determined by its amino acid sequence and the noncovalent interactions that stabilize its structure. In many cases, the conformation, and thus the function, of a protein can change when that protein associates noncovalently or covalently with other molecules or ions. Through these associations, the function — also called the activity — of a protein can be regulated (e.g., turned “on” or “off,” or “up” or “down”) to permit cells to adapt to changing conditions. Altered conditions include changes in nutrient availability (Chapter 21), hormonal signaling (Chapters 15 and 16), communication with other cells (Chapters 20 and 23), the developmental state of the organism, and the presence of pathogens (Chapter 24), along with many other factors. How many proteins are there in a typical eukaryotic cell? We can calculate that there are about 7.9×109 protein molecules in a mammalian hepatocyte (a liver cell). (This calculation is worked out in Section 3.1.) It is estimated that a hepatocyte contains about 10,000 different proteins; thus each cell, on average, contains close to a million molecules of each type of protein. In reality though, the abundances of different proteins vary widely, from the quite rare insulin-binding receptor protein (2×104 molecules per cell) to the structural protein actin (5×108 molecules per cell). Every cell closely regulates the abundance of each of its proteins so that they are present in the appropriate quantities to support the cell’s needs at any given time. We will learn more about the mechanisms used by cells to regulate protein levels later in this chapter and in Chapters 8 and 9. Because of their many different shapes and chemical properties, proteins can perform a dazzling array of functions inside and outside cells that either are essential for life or provide a selective evolutionary advantage to the cell or organism that contains them. It is, therefore, not surprising that characterizing the structures and activities of proteins — and how these change in response to regulation — is a fundamental prerequisite for understanding how cells work. Much of this textbook is devoted to examining how proteins act with one another or with other types of molecules (e.g., DNA) to allow cells to live and function properly. In this chapter we emphasize the basic principles underlying the structures and functions of proteins and the regulation of their activity. We also look at some of the many methods used to study them. Although their structures are diverse, most proteins can be grouped into a few broad functional classes. Structural proteins, for example, determine the shapes of cells and their extracellular environments and serve as guide wires or rails to direct the intracellular movement of molecules and organelles. Structural proteins are usually formed by the assembly of multiple protein subunits into very large and often very long structures. Scaffold proteins bring other proteins together into ordered arrays to perform specific functions more efficiently than those proteins would if they were not assembled together. Enzymes are proteins that catalyze chemical reactions (sometimes called molecular transformations). These molecular transformations are the fundamental activities in metabolic pathways (Chapter 12). They can also result in the modification of proteins to alter their activities (e.g., phosphorylation and dephosphorylation to activate and inactivate other proteins). Membrane transport proteins are embedded in cellular membranes and permit the flow of ions and molecules across the membranes. Regulatory proteins act as signals, sensors, and switches to control activities of cells by altering the functions of other proteins and genes. Regulatory proteins include signaling proteins, such as some hormones and cell-surface receptors that transmit extracellular signals to the cell interior. Motor proteins are responsible for the movement of other proteins, organelles, and cells — even whole organisms. There are additional proteins that don’t fit neatly into one of these classes. For example, fish that live in frigid waters — the Antarctic notothenioids and Arctic cods — have antifreeze proteins in their circulatory systems to prevent water crystallization. Many proteins are members of more than one class, such as some cell-surface signaling receptors that are enzymes and regulatory proteins because they transmit signals from outside to inside cells by catalyzing chemical reactions. To accomplish their diverse missions efficiently, some proteins assemble into very large complexes, often called molecular machines. How do proteins perform so many diverse functions? They do so by exploiting three simple mechanisms. The most fundamental is binding: proteins bind to one another, to other macromolecules such as DNA, and to small molecules and ions. Binding is based on molecular complementarity between a protein and its binding partner, as described in Chapter 2. A second mechanism is enzymatic catalysis. In enzymes, the proper conformation of the protein will place some amino acid side chains and some carboxyl and amino groups of its backbone into positions that permit the catalysis of covalent bond rearrangements in other molecules (called substrates of the enzyme). A third mechanism is regulation of protein activity. Typically, a protein’s shape or activity is altered through noncovalent (binding) or covalent (catalysis) association of molecules or ions with the protein. In many cases, this binding or catalysis induces a conformational change in the protein and that influences its activity. A complete understanding of how proteins permit cells to live and thrive requires that we identify and characterize all of the proteins used by a cell. In a sense, molecular cell biologists want to compile a complete protein “parts list” and then construct a “user’s manual” that describes how these proteins work. Compiling a comprehensive inventory of proteins has become feasible in recent years thanks to the sequenced genomes — complete sets of genes — of many organisms. From a computer analysis of a genome’s sequence, researchers can deduce the amino acid sequences and approximate number of different types of proteins the genome encodes (see Chapter 6). It is also possible to determine the sequences and relative amounts of a substantial fraction of the messenger RNAs (mRNAs) in individual cells or most of the mRNAs (the transcriptome) from a collection of similar cells and thus deduce what subset of proteins encoded in the genome is made (expressed) in a given type of cell. DNA and mRNA sequencing are indirect methods of characterizing the potential collection of proteins in cells. There are also methods that directly measure the collection of proteins in samples of cells, which we discuss in Sections 3.5 and 3.6. The term proteome was coined to refer to the entire protein complement of an organism, organ, or a particular type of cell within an organism. The human genome contains about 21,500 genes that encode proteins. However, variations in mRNA production, such as alternative splicing (see Chapter 9), and more than a hundred types of protein modifications may generate hundreds of thousands of distinct types of human proteins. By comparing the sequences and structures of proteins of unknown function with those of proteins of known function, scientists can often deduce much about what the unknown proteins do. In the past, a protein’s function was characterized by genetic, biochemical, or physiological methods before the particular protein had been identified. In the modern genomic and proteomic era, a protein is often identified before its function is determined. In this chapter, we begin our study of how the structure of a protein gives rise to its function, a theme that recurs throughout this book (Figure 3-1). In Section 3.1, we examine how linear chains of amino acids are arranged in a three-dimensional structural hierarchy. The next section discusses how proteins fold into these structures. We then turn to protein function, focusing on enzymes, those proteins that catalyze chemical reactions. Our focus on protein structure and function is part of a broader interest in cell biology in the principles relating biological structure and function. These principles were initially formulated by biologists Johann von Goethe (1749–1832), Ernst Haeckel (1834–1919), and D’Arcy Thompson (1860–1948), whose work has been widely influential in biology and beyond. Indeed, their ideas greatly influenced the school of organic architecture pioneered in the early twentieth century that is epitomized by the dicta “form follows function” (Louis Sullivan) and “form is function” (Frank Lloyd Wright). After considering protein structure and function, we turn to various mechanisms that cells use to control the activities and life spans of proteins. Particularly important control mechanisms are allosteric effector-binding, covalent phosphorylation, and ubiquitinylation of proteins. The chapter concludes with a discussion of commonly used techniques for identifying, isolating, and characterizing proteins, and a discussion of the burgeoning field of proteomics. FIGURE 3-1 Overview of protein structure and function. (a) Proteins have a hierarchical structure. A polypeptide’s linear sequence of amino acids linked by peptide bonds (primary structure) folds into local helices or sheets (secondary structure) that pack into a complex three-dimensional shape (tertiary structure). Some individual polypeptides associate into multichain complexes (quaternary structure), which in some cases can be very large, consisting of tens to hundreds of subunits (supramolecular complexes). (b) Proteins perform numerous functions, including organizing the genome, organelles, cytoplasm, protein complexes, and membranes in three-dimensional space (structure); monitoring the environment and transmitting information (signaling); moving small molecules and ions across membranes (transport); catalyzing chemical reactions that result in the molecular transformation of one molecule into another; and generating force for movement (via motor proteins). These functions and others arise from mechanisms involving binding, catalysis, and regulation. In illustration A, the primary (sequence) structure leads to the secondary (local folding and hydrogen bonding) structure that further leads to tertiary (overall conformation) structure. This leads to a quaternary (multimeric) structure which further leads to a supramolecular (large-scale assembly) structure. In illustration B, the tertiary, quaternary, and supramolecular structures lead to several functions like binding, catalysis, and regulation. These further help in signaling, transport, molecular transformation, movement, and structure.