In vivo and in vitro studies of transmembrane β-strand deletion, insertion or substitution mutants of the Escherichia coli K-12 maltoporin

Abstract

Lame of Escherichia coli K12, also called maltoporin, is an outer membrane protein, which specifically facilitates the diffusion of maltose and maltodextrin through the bacterial outer membrane. Each monomer is composed of an 18-stranded antiparallel β-barrel. In the present work, on the basis of the known X-ray structure of LamB, the effects of modifications of the β-barrel domain of maltoporin were studied in vivo and in vitro. We show that: (i) the substitution of the pair of strands β13-β14 of the E. coli maltoporin with the corresponding pair of strands from the functionally related maltoporin of Salmonella typhimurium yielded a protein active in vivo and in vitro; and (ii) the removal of one pair of β-strands (deletion β13-β14) from the E. coli maltoporin, or its replacement by a pair of strands from the general porin OmpF of E. coli, leads to recombinant proteins that lost in vivo maltoporin activities but still kept channel formation and carbohydrate binding in vitro. We also inserted into deletion β13-β14 the portion of the E. coli LamB protein comprising strands β13 to β16. This resulted in a protein expected to have 20 β-strands and which completely lost all LamB-specific activities in vivo and in vitro.