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X-ray structural determination and biophysical characterization of HemAT, a chemotaxis receptor from Bacillus subtilis

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dc.contributor.advisor Phillips, George N., Jr.
dc.creator Zhang, Wei
dc.date.accessioned 2009-06-04T08:32:00Z
dc.date.available 2009-06-04T08:32:00Z
dc.date.issued 2004
dc.identifier.uri http://hdl.handle.net/1911/18725
dc.description.abstract The heme-based aerotaxis transducer (HemAT) from B. subtilis is a heme-containing protein and functions as an oxygen sensor. It can detect oxygen and transmit the signal generated from oxygen binding to regulatory proteins through its putative methyl-accepting chemotactic domain. Through other components, the signaling information is transferred to motor proteins, which control the direction of rotation of flagella and in turn lead to changes in the swimming behavior of bacteria. There is a great deal of information known about chemotaxis signaling transduction for Escherichia coli and Salmonella typhimurium. However, the detailed molecular mechanism of chemotaxis of Bacillus subtilis is in a sense reversed, because attractant binding to chemotactic receptors strengthens the activity of the downstream histidine kinase, instead of inhibiting reaction in Escherichia coli and Salmonella typhimurium. Multiple-wavelength anomalous dispersion (MAD) data were collected from crystals of HemAT using the intrinsic anomalous scatterer, iron, with synchrotron radiation. Three wavelength iron MAD data were collected to 2.8A resolution. The native data set was collected to 2.15A resolution. The crystallographic analysis reveals that the crystal belongs to P21212 1 space group with the cell dimension a = 50.00A, b = 80.12A, c = 85.95A. There are two molecules in one asymmetric unit with 40% solvent content. I have determined the crystal structures of the HemAT sensor domain in liganded and unliganded forms at resolutions of 2.15A and 2.7A. The structures show that the HemAT sensor domain is a dimeric protein with one heme group in each subunit. The structure of liganded form of HemAT sensor domain reveals a more symmetrical organization than that of the unliganded form. Tyrosine70 in one subunit shows distinct conformations in the liganded and unliganded structures. Our study suggests that disruption of HemAT symmetry plays an important role in initiating the chemotaxis signaling transduction pathway. Our kinetic and thermodynamic studies of ligand binding suggest that HemAT may employ negative cooperativity for detecting external ligand in the signal transduction. The sensor domain provides the structural evidence for such a molecular mechanism.
dc.format.extent 167 p.
dc.format.mimetype application/pdf
dc.language.iso eng
dc.subject Chemistry, Biochemistry
Biophysics, General
dc.title X-ray structural determination and biophysical characterization of HemAT, a chemotaxis receptor from Bacillus subtilis
dc.type.genre Thesis
dc.type.material Text
thesis.degree.discipline Chemistry
thesis.degree.discipline Biophysics
thesis.degree.grantor Rice University
thesis.degree.level Doctoral
thesis.degree.name Doctor of Philosophy
dc.identifier.citation Zhang, Wei. (2004) "X-ray structural determination and biophysical characterization of HemAT, a chemotaxis receptor from Bacillus subtilis." Doctoral Thesis, Rice University. http://hdl.handle.net/1911/18725.

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