Strategies and considerations for protein purifications.

Abstract Prior to embarking upon the purification of a protein, one should begin by considering what the protein is to be used for. In particular, how much of the protein is needed, what should be its state of purity, and must it be folded correctly and associated with various other peptides or cofactors. Using such criteria, an appropriate assay should be chosen and a procedure be planned taking into account the source of the protein, how it is to be extracted from the source, and what agents the protein ought to be exposed to or ultimately be stored in. One is often surprised at the time necessary to develop an appropriate protein purification procedure relative to the time required to clone a gene or to accumulate information with the purified protein. There are an overwhelming number of options for protein purification steps, so forethought is necessary to expedite the tedious job of developing the purification scheme, or to avoid having to redesign it upon attempting to use the protein. This chapter points out general considerations to be undertaken in designing, organizing, and executing the purification, while subsequent chapters of this volume supply more specific options and technical details.