EXPRESSION OF LARGE AMOUNTS OF NEISSERIAL PORIN PROTEINS IN ESCHERICHIA-COLI AND REFOLDING OF THE PROTEINS INTO NATIVE TRIMERS

Porins from different neisserial strains and species have been shown t o have differences in both primary amino acid sequence and biophysical characteristics as observed by functional assays. A closer examinatio n of how the changes in the primary amino acid sequence of Neisseria p orin molecules correlate with these observed biophysical changes has b een impeded by the inability to easily manipulate the cloned porin gen es by modern molecular techniques and then obtain enough of the expres sed modified porin protein to purify and use in these biophysical func tional assays. In this report, we describe a method by which the genes encoding three different porin proteins, lacking their neisserial pro moter and signal sequences, were cloned into an expression plasmid and transformed into Escherichia coli. Upon induction, large amounts of t he porin proteins were produced. The expressed porin proteins were the n manipulated to regenerate their native trimer structure and purified by standard protein chemistry. Sufficient purified recombinant porin protein was obtained for further antigenic as well as biophysical char acterization. This sets the stage for the biophysical characterization of these neisserial porin proteins in more detail.