| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
Biophys J, July 2002, p. 309-321, Vol. 83, No. 1
Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum) der Universität Würzburg, Am Hubland, D-97074 Würzburg, Germany
The 3-D structure of the maltooligosaccharide-specific
LamB channel of Escherichia coli (also called
maltoporin) is known from x-ray crystallography. The central
constriction of the channel formed by the external loop 3 is controlled
by tyrosine 118. Y118 was replaced by site-directed mutagenesis by 10 other amino acids (alanine (A), isoleucine (I), asparagine (N), serine
(S), cysteine (C), aspartic acid (D), arginine (R), histidine (H),
phenylalanine (F), and tryptophan (W)) including neutral ones,
negatively and positively charged amino acids to study the effect of
their size, their hydrophobicity index, and their charge on maltose and
maltooligosaccharide binding to LamB. The mutants were reconstituted
into lipid bilayer membranes and the stability constants for binding of
maltose, maltotriose, maltopentaose, and maltoheptaose to the channel
were measured using titration experiments. The mutation of Y118 to any
other non-aromatic amino acid led to a substantial decrease of the
stability constant of binding by factors between about two and six. The
highest effect was observed for the mutant Y118A. Replacement of Y118
by the two other aromatic amino acids, phenylalanine (F) and tryptophan
(W), resulted in a substantial increase of the stability constant
maximally by a factor of almost 400 for the Y118W mutant. The
carbohydrate-induced block of the channel function was used for the
study of current noise through the different mutant LamB channels. The
analysis of the power density spectra allowed the evaluation of the on-
and off-rate constants (k1 and k
1) of sugar binding. The results suggest
that both rate constants were affected by the mutations. For most
mutants, with the exception of Y118F and Y118W,
k1 decreased and
k1 increased, whereas the opposite was
found for the aromatic amino acid mutants. The results suggest that
tyrosine 118 has a crucial effect on carbohydrate transport through LamB.
Biophys J, July 2002, p. 309-321, Vol. 83, No. 1
© 2002 by the Biophysical Society 0006-3495/02/07/309/13 $2.00
This article has been cited by other articles:
 |
|
 |
|
  F. Orlik, B. Schiffler, and R. Benz Anthrax Toxin Protective Antigen: Inhibition of Channel Function by Chloroquine and Related Compounds and Study of Binding Kinetics Using the Current Noise Analysis Biophys. J., March 1, 2005; 88(3): 1715 - 1724. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Danelon, T. Brando, and M. Winterhalter Probing the Orientation of Reconstituted Maltoporin Channels at the Single-protein Level J. Biol. Chem., September 12, 2003; 278(37): 35542 - 35551. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Orlik, C. Andersen, C. Danelon, M. Winterhalter, M. Pajatsch, A. Bock, and R. Benz CymA of Klebsiella oxytoca Outer Membrane: Binding of Cyclodextrins and Study of the Current Noise of the Open Channel Biophys. J., August 1, 2003; 85(2): 876 - 885. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Andersen, B. Schiffler, A. Charbit, and R. Benz pH-induced Collapse of the Extracellular Loops Closes Escherichia coli Maltoporin and Allows the Study of Asymmetric Sugar Binding J. Biol. Chem., October 25, 2002; 277(44): 41318 - 41325. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
