Supramolecular Protein Chemistry

Assembly, Architecture and Application

Peter B Crowley, Peter B Crowley

• Publisher: Royal Society of Chemistry
• Year: 2020
• ISBN:
  9781788019804
• Edition: 1
• Topic: Physical Sciences
• Subtopic: Physical & Theoretical Chemistry

Building on decades of "host-guest" research, recent years have seen a surge of activity in water-soluble supramolecular receptors for protein recognition and assembly. Progress has been particularly rich in the area of calixarenes, cucurbiturils and molecular tweezers. Emerging applications include controlled protein assembly in solution, crystal engineering, supramolecular control of catalysis (both in vitro and in vivo), as well as novel mechanisms of protein-interaction inhibition with relevance to amyloids and disease. One challenge at the interface of supramolecular chemistry and protein science is to increase interaction and collaboration between chemists and biochemists/structural biologists.This book addresses the exciting interface of supramolecular chemistry and protein science. Chapters cover supramolecular approaches to protein recognition, assembly and regulation. Principles outlined will highlight the opportunities that are readily accessible to collaborating chemists and biochemists, enriching the breadth and scope of this multidisciplinary field. Supramolecular Protein Chemistry will be of particular interest to graduate students and researchers working in supramolecular chemistry, protein science, self-assembly, biomaterials, biomedicine and biotechnology.

CHAPTER 1

Supramolecular Approaches to Protein Recognition

JOSEPH W. MEISEL^a, ^b AND ANDREW D. HAMILTON^*a

^a Department of Chemistry, New York University, 100 Washington Square East, New York, NY 10012, USA, ^b Department of Chemistry, The George Washington University, 800 22nd Street NW, Washington, DC 20052, USA
^*E-mail: [email protected]

This chapter begins with an account of the early developments in protein chemistry and supramolecular chemistry. We have attempted to summarize the seminal achievements in each of these vast fields, highlighting how one has informed the other over the past century. We focus on the central topic of molecular recognition and noncovalent interactions. Classic examples of molecular recognition in Nature (as evidenced by X-ray crystallography) and the corresponding motifs developed by supramolecular chemists are described. Synthetic scaffolds that mimic peptide and protein structures are also presented in the context of interaction inhibitors. By bringing together these illustrious examples we hope to encourage new collaborations between supramolecular chemists and biomedical scientists.

1.1 Introduction

Proteins are biological macromolecules that are ubiquitous in living systems. They play a role in nearly every aspect of cellular function, often carrying out their multifarious roles through noncovalent interactions with other molecules. As protein activity is so frequently mediated by assembly with other structures, it is critical to understand the forces that determine their supramolecular chemistry. Through this perspective, we may better comprehend the chemical basis for protein function and can more readily manipulate their activity. An important objective of this book is to stimulate greater collaboration between supramolecular chemists and biomedical scientists studying protein structure and function. As the authors believe that common foundational knowledge is critical to meaningful multidisciplinary research, we have elected to begin this chapter with a historical introduction to both supramolecular and protein chemistry fields. Where possible, we have referenced the original literature both for posterity and for the benefit of the unique perspective these primary accounts provide.

1.2 Early Protein Chemistry

Long before the composition of proteins was known, their biological importance was already apparent to the early 19th century chemists who studied them. To reflect this sentiment, the term protein was coined by Gerardus Johannes Mulder as a derivative of the Greek word πρωτειoς (proteios), meaning first place or primary. Mulder was inspired by Jöns Jacob Berzelius who proposed the word in an 1838 letter to Mulder, stating that protein “appears to be the primitive or principal substance of animal nutrition”.¹ A major focus of Berzelius, Mulder, and other early protein chemists was to apply increasingly sophisticated combustion analysis methods to determine the elemental constitution of purified protein. They observed similar empirical formulas for protein extracted from different plants and animals. This observation suggested that protein was a fundamental component of all living organisms. Furthermore, their experiments implied that proteins were macromolecules as their molecular weights were much larger than any other substance known at that time. Although the complete picture of protein structure would elude scientists for another century, the peculiar solubility properties were conspicuous. Unlike most compounds familiar to chemists at that time, proteins are less soluble at higher temperatures and are sensitive to the presence of different salts. Franz Hofmeister and co-workers carried out systematic investigations on the influence of salts on the solubility of proteins isolated from egg white and blood serum.² They found that certain ions would increase protein solubility (salting in) and others would decrease solubility (salting out). The rank order of salts is known as the Hofmeister series and it continues to reveal fundamental properties of supramolecular interactions in water³ and to have practical applications in industry.⁴ Another major contribution to protein chemistry by Hofmeister was the proposal that proteins comprise amino acids linked together.⁵ Emil Fischer made this discovery independently⁶ and proposed the term peptide for the bond that links amino acids. Coincidentally, Fischer and Hofmeister both reported their findings at the same Karlsbad conference in 1902.

Despite significant work supporting the polypeptide theory of protein composition, the molecular nature of proteins was not universally accepted. Many respected chemists simply resisted the possibility that such large molecules could exist. The most vocal opposition came from proponents of the colloidal theory,⁷ who posited that protein was a suspension of charged particles rather than discrete molecular entities. The colloid chemists' reductionist approach was an oversimplification; they viewed proteins as hard spheres and ellipsoids. Nevertheless, their quantitative physicochemical methodology provided many insights into protein behaviour through measurements of acid-base equilibria, osmotic pressure, and hydrodynamic properties.⁸ While the molecular and colloidal camps disagreed on the structure of proteins, both parties could agree that proteins were large. It was not until the 1950s that the molecularity of proteins was fully accepted upon publication of the complete sequence of insulin by Frederick Sanger.^9,10 Surprisingly, this result came three decades after Svedberg's ultracentrifugation experiments proved that haemoglobin was a tetramer composed of four proteins weighing 16.7 kDa each.¹¹ Thus, the supramolecular nature of protein quaternary structures was apparent long before a primary structure was known.

In many cases, the biochemical function of proteins was studied before their molecular structure was known. This characteristic is especially apparent in enzymology, much of which evolved from observations on the catalytic digestion of foodstuffs. For example, malting and mashing are ancient processes that use seed enzymes to convert the starch in raw grain into oligosaccharides, primarily for sugar production or fermentation. In 1833, Payen and Persoz reported the extraction of the enzyme diastase from malt.¹² The activities of diastase and the digestive enzyme pepsin, discovered by Theodor Schwann in 1836,¹³ were the first examples of biological processes occurring outside of organisms. Although the enzymatic material clearly comprised protein, it was not known if the catalytic action was due to some other compound and the protein was simply an impurity. Due to the difficulties in protein purification, it took nearly a century for enzymes to be definitively recognized as proteins, with sceptics finally capitulating when Sumner crystallized urease¹⁴ and Northrup crystallized pepsin.¹⁵ The two shared the 1946 Nobel Prize with the virologist Wendell Meredith Stanley for this work.

The catalytic property of enzymes is only one aspect of their impressive chemical activity. Their ability to act only on a particular substrate and to engender a singular product, i.e. their specificity, is a marvel. Fourteen years before he described the polypeptide nature of protein structure, Fischer proposed his famous lock and key (Schloss und Schlüssel) model to explain enzyme specificity.¹⁶ Fischer was studying the action of enzymes on carbohydrate stereoisomer inversion. The lock and key model was revolutionary for several reasons. First, the model implied that the mechanism of enzyme action is through the physical combination of enzyme and substrate, while earlier theories attributed a more nebulous ‘vital force’. Further, the model assimilated the organic chemists' burgeoning three-dimensional understanding of molecular structure, sterics, and stereochemistry with the less-understood structure of enzymes. Thus, by relating function to structure, the lock and key model beckoned further research into protein composition and conformation. This impetus to ‘think three-dimensionally’ is shared among structural biologists and supramolecular chemists alike.

Contemporaneous with advances in enzymology and organic chemistry, immunology was another emerging discipline that would contribute greatly to protein chemistry. Although inoculation existed for centuries before Edward Jenner's smallpox vaccine was introduced in 1798, it was nearly a century later when immunology began to enter the realm of chemistry. In 1890, Emil Behring and Kitasato Shibasaburō derived tetanus and diphtheria antitoxins from the serum of immunized animals.¹⁷ Their excitement is palpable as the article concludes with the quotation of Goethe: “Blut is ein ganz besonderer Saft” (blood is a very special fluid). The shift toward a molecular perspective is evident when Paul Ehrlich proposed his side-chain theory to describe the action and production of antibodies several years later. At the same time, Gruber and Durham discovered the agglutination reaction and Kraus developed the precipitin reactions.¹⁸ Both tests are used to quantify antibody titres in blood by the addition of varying amounts of antigen to serum. If antibodies are present and are specific to the antigen (and multivalent), their supramolecular interactions induce cross-linking observable as precipitation. Parallel to the acceleration of enzymology, which was catalysed by the food and chemical industries, medicine bolstered immunology research. Precipitin-based testing became a standard diagnostic assay for infectious disease and antibody serum therapies were successful treatments thereof.

Chemists and physiologists continued to refine their theories to explain biological phenomena. However, the limitations in the purification and chemical characterization of protein remained. As a result, in 1905 John Langley was able to obtain pure nicotine and de...