APPLICATION OF GENE-TRANSFER TECHNOLOGIES TO THE PRODUCTION OF ENZYMEREFERENCE MATERIALS - EXAMPLE OF GAMMA-GLUTAMYL-TRANSFERASE

Protein reference materials are traditionally prepared by purification from mammalian or human tissues. The supply of these tissues is limit ed; consequently, there is a growing need for applied molecular and ce llular biology technologies for the production of human recombinant pr oteins. This is especially true when only small amounts of the protein s are available in the tissues. We review the current knowledge necess ary for high-level production of such proteins in different heterologo us expression systems, using our data on gamma-glutamyltransferase (EC 2.3.2.2) as an example. We describe the steps required to achieve the expression of enzymes and other proteins in Escherichia coli, yeast, or mammalian cells. We list many of the problems investigators may fac e in preparing recombinant proteins, and provide information on select ing the most appropriate system as well as the most favorable experime ntal conditions. Depending on the expression system, recombinant prote ins can potentially be obtained for most, if not all, enzymes of inter est in clinical chemistry, and such proteins should possess characteri stics very similar to those of the corresponding human native proteins . Studies suggest that these products can be used as reference materia ls in clinical chemistry laboratories.