| Autor: |
Jordy, Malte, Andersen, Christian, Schülein, Katrin, Ferenci, Tom, Benz, Roland |
| Zdroj: |
JMB Online (Journal of Molecular Biology); June 21, 1996, Vol. 259 Issue: 4 p666-678, 13p |
| Abstrakt: |
Two LamB (maltoporin) point mutants ofEscherichia coli(R8H and Y118F) and wild-type LamB ofSalmonella typhimuriumwere reconstituted into artificial lipid bilayer membranes. Ion transport through wild-type LamB ofS. typhimuriumand the LamB mutants was inhibited by the addition of carbohydrates of maltose and maltooligosaccharide type in a dose-depen dent fashion. The sugar-induced block of the channel function could be used for the study of current noise through the different wild-type and mutant LamB-channels. The analysis of the power density spectra allowed the evaluation of the on and off-reactions (klandk−1) of sugar-binding to the binding site inside the channels. Wild-type LamB ofS. typhimuriumhad approximately the same sugar-binding kinetics as has been observed for LamB ofE. coli. The results suggest that the binding site inside the channel interacts with a maximum of three glucose residues within the maltooligosaccharides. The LamB mutants R8H and Y118F showed kinetics for sugar binding substantially different from that of wild-type LamB. In particular, the on-rate,kl, for the binding of different sugars of the maltooligosaccharide series to the mutant R8H was approximately 500-times smaller than for wild-type LamB, which resulted in a substantially smaller stability constant of sugar binding to the channel. Similarly, the off-rate constant,k−1, for sugar binding to the mutant Y118F decreased about 20-fold, which led to a strong increase of the affinity of carbohydrates to the site. The role of the amino residues acid R8 and Y118 in the transport of maltose and maltooligosaccharides through LamB-channels is discussed on the basis of the net flux of sugars through the channels. |
| Databáze: |
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