POSTERS - BLAST X - University of Utah
POSTERS - BLAST X - University of Utah
POSTERS - BLAST X - University of Utah
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<strong>BLAST</strong> X Wed. Morning Session<br />
THE TM2-HAMP CONNECTION<br />
Gus A. Wright, Rachel L. Crowder and Michael D. Manson<br />
Department <strong>of</strong> Biology, Texas A&M <strong>University</strong>, College Station, TX 77843<br />
The HAMP domain—a conserved protein motif in most Histidine kinases, Adenylate<br />
cyclases, Methyl-accepting chemotaxis proteins, and Phosphatases—typically conforms to a<br />
helix–connector-helix domain architecture. Recently, an NMR solution structure was determined<br />
for the Af1503 HAMP from the archaebacteria Archeoglobus fulgidis (Hulko, et al, Cell 126: 929-<br />
940, 2006). This structure revealed that the HAMP domain is a parallel four-helix bundle with a<br />
knob-on-knob packing. Similar structures <strong>of</strong> HAMP domains are proposed to exist in all 5<br />
chemoreceptors <strong>of</strong> Escherichia coli, including the aspartate chemoreceptor Tar. The HAMP<br />
domain <strong>of</strong> Tar receives information from TM2 and regulates the transmission <strong>of</strong> the signal to the<br />
kinase signaling domain. Mutations were introduced into the TM2-HAMP connector region <strong>of</strong><br />
Tar to determine if manipulating the input signal into the HAMP affects the output signal to the<br />
signaling domain. MLLT residues between R214 and P219 were deleted (-4 through -1), and<br />
additional LLT tandem repeats were added up to 8 residues after P219 (+1 through +8).<br />
Aspartate sensitivity, rotational bias, mean reversal frequency, and in vivo methylation were<br />
measured for these mutants. The results suggest that adding helical turns in this region<br />
destabilizes, or “relaxes”, the HAMP/signaling domains; while, removing helical turns from this<br />
region stabilizes, or “tightens”, the HAMP/signaling domains. In order to determine if increasing<br />
flexibility <strong>of</strong> the TM2/HAMP connector affects the output signal, a second set <strong>of</strong> mutants were<br />
constructed to replace the MLLT region with four tandem glycine residues (4G). Glycine<br />
residues were then subtracted (-4G through -1G) and added (+1G through +5G). Data collected<br />
from this experiment suggests that increasing the flexibility <strong>of</strong> this region dampens the input<br />
signal from TM2, in addition to destabilizing the HAMP/signaling domains.<br />
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