This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.The bacterial second messenger bis-(3 '-5 ')-cyclic dimeric guanosine monophosphate (c-di-GMP) regulates complex biological processes such as cell motility, biofilm formation and virulence. Protein domains catalyzing synthesis and degradation of c-di-GMP, GGDEF and EAL domains, respectively, have been identified in large numbers in almost every microbial genome sequenced to date. Our understanding regarding function and regulation of these enzymes is only in its infancy. Using structural biology in combination with enzymatic assays, we hope to decipher the regulatory mechanisms and modes of operation of key enzymes controlling biofilm formation and cytotoxicity, using the opportunistic pathogen Pseudomonas aeruginosa as a model system. Results will have widespread applications such as the development of novel antibiotics for the treatment of infectious diseases, optimization of biotechnological processes, or the development of novel biomedical therapies. In more general terms, we are interested in the intra- and intermolecular regulatory principles in multi-domain proteins, including allosteric and feedback control, fundamental questions in signal transduction and enzyme regulation. We obtained crystals for three proteins containing EAL-, GGDEF- or tandem-domains in various conditions. Preliminary X-ray diffraction experiments performed at CHESS, beamline F1 show promising results. Crystals diffract X-rays to a maximum resolution of 10 angstrom but we are confident that improved crystallization and freezing conditions will increase the diffraction limit. Given the small size of some of the crystals, we require a bright light source for crystal screening and data collection.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR001646-25
Application #
7598564
Study Section
Special Emphasis Panel (ZRG1-BBCA (40))
Project Start
2007-07-01
Project End
2008-06-30
Budget Start
2007-07-01
Budget End
2008-06-30
Support Year
25
Fiscal Year
2007
Total Cost
$37,720
Indirect Cost
Name
Cornell University
Department
Physics
Type
Schools of Arts and Sciences
DUNS #
872612445
City
Ithaca
State
NY
Country
United States
Zip Code
14850
Chen, Yu Seby; Kozlov, Guennadi; Fakih, Rayan et al. (2018) The cyclic nucleotide-binding homology domain of the integral membrane protein CNNM mediates dimerization and is required for Mg2+ efflux activity. J Biol Chem 293:19998-20007
Kozlov, Guennadi; Wong, Kathy; Gehring, Kalle (2018) Crystal structure of the Legionella effector Lem22. Proteins 86:263-267
Ménade, Marie; Kozlov, Guennadi; Trempe, Jean-François et al. (2018) Structures of ubiquitin-like (Ubl) and Hsp90-like domains of sacsin provide insight into pathological mutations. J Biol Chem 293:12832-12842
Xu, Jie; Kozlov, Guennadi; McPherson, Peter S et al. (2018) A PH-like domain of the Rab12 guanine nucleotide exchange factor DENND3 binds actin and is required for autophagy. J Biol Chem 293:4566-4574
Dean, Dexter N; Rana, Pratip; Campbell, Ryan P et al. (2018) Propagation of an A? Dodecamer Strain Involves a Three-Step Mechanism and a Key Intermediate. Biophys J 114:539-549
Xu, Caishuang; Kozlov, Guennadi; Wong, Kathy et al. (2016) Crystal Structure of the Salmonella Typhimurium Effector GtgE. PLoS One 11:e0166643
Cogliati, Massimo; Zani, Alberto; Rickerts, Volker et al. (2016) Multilocus sequence typing analysis reveals that Cryptococcus neoformans var. neoformans is a recombinant population. Fungal Genet Biol 87:22-9
Oot, Rebecca A; Kane, Patricia M; Berry, Edward A et al. (2016) Crystal structure of yeast V1-ATPase in the autoinhibited state. EMBO J 35:1694-706
Lucido, Michael J; Orlando, Benjamin J; Vecchio, Alex J et al. (2016) Crystal Structure of Aspirin-Acetylated Human Cyclooxygenase-2: Insight into the Formation of Products with Reversed Stereochemistry. Biochemistry 55:1226-38
Bauman, Joseph D; Harrison, Jerry Joe E K; Arnold, Eddy (2016) Rapid experimental SAD phasing and hot-spot identification with halogenated fragments. IUCrJ 3:51-60

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