POSTERS - BLAST X - University of Utah

BLAST X Wed. Morning Session
A NOVEL AMINO ACID BINDING STRUCTURE IN BACTERIAL CHEMOTAXIS
George D. Glekas and George W. Ordal
Department of Biochemistry, University of Illinois at Urbana-Champaign
409 MSB, Urbana, IL 61801
Simple flagellated bacteria, such as Bacillus subtilis, possess the ability to sense their
environment and move to more favorable conditions, where there are, for instance, more
nutrients like amino acids, by the process of chemotaxis. The initial stage is binding of an
amino acid by receptors on the outside of the cell. This binding causes conformational changes
that affects the activity of enzymes on the inside of the cell and alters the movement of the
bacteria. The main paradigm for understanding these events is the bacterium Escherichia coli.
However, we have discovered that, in fact, the mechanism used by E. coli is not used by most
bacteria and that the mechanism used by the distantly related bacterium B. subtilis is more likely
to be the general mechanism. Unlike in E. coli, where binding of attractant causes a shift of the
receptor polypeptide that goes from the outside of the cell to the cell interior toward the cell
interior, attractant causes rotational movement of the receptors without any comparable interior
shifting
We seek to understand how this rotational movement occurs in the asparagine receptor
McpB. To do this, we have discovered that the most likely conformation of the exterior part of
the receptor is far different from that in the E. coli receptor. Molecular and homology modeling
of the McpB sensing domain has led to a structural model that reveals a dual PAS domain
structure similar to the crystal structure of the LuxQ sensor. PAS domains are known to be
conserved structures capable of binding a great many “small” molecules. Further mutagenetic
analyses of putative asparagine-binding residues have not only confirmed the validity of the
structural model, they have revealed certain residues that greatly effect chemo-attractant
binding. Using both in vivo chemotactic assays and in vitro isothermal titration calorimetry
performed on purified mutant receptor exterior regions, three residues, all in the upper PAS
domain, have been shown to lower the affinity of the McpB receptor for asparagine. Mutations
in the lower PAS domain show no such effect. Further structural and mutagenetic studies have
shown a similar dual PAS architecture in the B. subtilis proline receptor McpC, with similar
residues responsible for amino acid binding. Extensive homology modeling shows that eight of
the ten B. subtilis chemoreceptors have PAS domains in their sensing domains.
We are now in the process of modeling the consequences of binding attractant at these
residues on the expected structure of the receptor.
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