Chapter 7b - Protein Methods

Summary

INTRODUCTION

In the post-genomic era, with sequencing of the entire human genome complete, there has been a great increase in our knowledge regarding the function of proteins encoded by the genome, the so-called proteome, and the role that aberrant protein expression and/or function plays in human disease pathogenesis. ‘Genomics’ refers to the comprehensive study of the function, interactions and dynamics of genes. In turn, ‘transcriptomics’, while having a similar objective, describes the function, interactions and dynamics of the genomic transcripts (i.e. mRNA, tRNA, rRNA and small RNAs such as miRNA and siRNA). ‘Proteomics’, by logical succession, describes the equivalent parameters for the complete set of proteins expressed by the genome through its transcripts. The entire set of proteins expressed by a particular cell/ organ ism constitutes its proteome. A number of variations on this theme are encountered increasingly in the literature. For instance, ‘metabolomics’refers specifically to the study of genes, transcripts and proteins involved in the regulation of metabolism.

In this chapter, we present a brief overview of some of the important tools that have been used to study the basic physicochemical properties of proteins, which have contributed significantly to developments in the field of proteomics. We also describe how these are increasingly being applied in the diagnosis and understanding of human disease.

PROTEIN BIOCHEMISTRY: BASIC CONCEPTS

The central dogma of molecular biology describes the process by which the genetic information contained within an organism's genes is translated into the functional components of life, proteins. This involves the produc tion of messenger RNA (mRNA) from the coding DNA sequence (transcription), which then serves as a template for protein synthesis (translation). In this manner, proteins are synthesised by polymerisation of the basic building blocks of all proteins, amino acids, in an order that is defined by the sequence of nucleotides in the coding DNA. It is the sequence in which these amino acids are incorporated into a protein molecule that is largely responsible for determining the threedimensional structure of the protein. This, in turn, is essentially what determines the biochemical activity of the protein and its role in the cell's life.