Application of gene transfer technologies to the production of enzyme reference materials: example of gamma-glutamyltransferase.
Protein reference materials are traditionally prepared by purification from mammalian or human tissues. The supply of these tissues is limited; consequently, there is a growing need for applied molecular and cellular biology technologies for the production of human recombinant proteins. This is especially true when only small amounts of the proteins are available in the tissues. We review the current knowledge necessary for high-level production of such proteins in different heterologous 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 face in preparing recombinant proteins, and provide information on selecting the most appropriate system as well as the most favorable experimental conditions. Depending on the expression system, recombinant proteins can potentially be obtained for most, if not all, enzymes of interest in clinical chemistry, and such proteins should possess characteristics very similar to those of the corresponding human native proteins. Studies suggest that these products can be used as reference materials in clinical chemistry laboratories.
Siest
G
G
Centre du Médicament, URA CNRS 597, Nancy, France.
Oster
T
T
Visvikis
A
A
Thioudellet
C
C
Angèle
C
C
Schiele
F
F
Wellman
M
M
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D011994:Recombinant Proteins / Q000096:biosynthesis; D012015:Reference Standards; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014162:Transfection; D005723:gamma-Glutamyltransferase / Q000096:biosynthesis / Q000235:genetics*
D011994:Recombinant Proteins; D005723:gamma-Glutamyltransferase