Expression of
recombinant proteins in bacteria has facilitated the characterization of many gene products. However, the biochemical characterization of
recombinant proteins is limited since the bacterially expressed
proteins are often synthesized as fusion
polypeptides. The presence of bacterial sequences in fusion
proteins further limits the use of these
proteins for generating
antibodies since the bacterial sequences are also antigenic. We describe two new bacterial expression vectors based on the pATH series of plasmids. These vectors were made by precisely deleting all of the trpE coding sequences found in pATH. The new vectors have enabled us to express eukaryotic genes as nonfusion
polypeptides. These altered plasmids can be used to insert any DNA sequence of interest through a multiple cloning site located just 3' of an ATG
start codon.
Protein expression is still under the control of the trp operon and is carried out at great efficiency when the bacteria are
tryptophan deprived. Studies presented here test the expression system with neurofilament subunits, NF-L and NF-H. Large amounts of recombinant nonfusion
proteins were produced. Also, a time course of induction shows that the production of the nonfusion
proteins was under the control of the trp operon which is readily inducible after
tryptophan starvation and addition of
indoleacrylic acid. These vectors may be useful for the overexpression of many
proteins in a form closely approximating their native state.