Domenic Castignetti
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Professor Ph.D., 1980, University of Massachusetts Microbial physiology and biochemistry Phone: 773.508.3638 Fax: 773.508.3646 E-Mail: dcastig@luc.edu |
RESEARCH INTERESTS
Siderophores are low molecular weight, avid ferric ion-binding compounds synthesized by fungi and bacteria. They sequester ferric ion from environments where its concentration is critically low. Without siderophores, microbes in such environments would cease growth due to lack of iron. Siderophores are employed by microbes to supply iron in environments such as soil, water and clinical infections. Previous research has established the synthesis and transport of siderophores and their role in the pathogenesis of the disease cystic fibrosis.We have noted that the mucoid bacterium Pseudomonas aeruginosa, the most prevalent infectious agent in cystic fibrosis, produces the same siderophores as its wild-type counter-part. We have also demonstrated that this bacterium uses siderophores to acquire ferric ion in the lung. Research is continuing to further understand the mechanism by which the bacterium establishes and maintains itself in CF-lung infections.
We also are examining the fate of siderophores in the environment. We have isolated a soil bacterium (a Rhizobium loti-like organism) that utilizes the siderophore deferrioxamine B (DFB) as its sole source of carbon and is thus able to return the molecule to the carbon cycle. Work has begun to elucidate the mechanism and the enzymes by which the bacterium is able to use DFB as a carbon source. The catalyst responsible for the initial breakdown of DFB, which we call DFB hydrolase, may be a single enzyme or an enzyme consortium. Studies, both biochemical and genetic, to elucidate which of these strategies the bacterium employs in its use of DFB, are underway in the laboratory.
REPRESENTATIVE PUBLICATIONS
Morton, J., K. Marsh, M. Frawley and D. Castignetti. 2007. The response of a siderophore-degrading bacterium (Mesorhizobium loti) to iron-deprivation: evidence of siderophore and iron-repressible protein synthesis. Adv. Biol. Res. (in press).Pierwola, A., T. Krupinski, P. Zalupski, M. Chiarelli and D. Castignetti. 2004. Degradation pathway and generation of monohydroxamic acids from the trihydroxamate siderophore deferrioxamine B. Appl. Environ. Microbiol. 70: 831-836.
Thupvong, T., A. Wiideman, D. Dunn, K. Oreschak, B. Jankowicz, J. Doering and D. Castignetti. 1999. Sequence heterogeneity of the ferripyoverdine uptake (fpvA), but not the ferric uptake regulator (fur), genes among strains of the fluorescent pseudomonads Pseudomonas aeruginosa, Pseudomonas aureofaciens, Pseudomonas fluorescens and Pseudomonas. BioMetals. 12:265-274.
Zaya, N., A. Roginsky, J. Williams and D. Castignetti. 1998. Evidence that a deferrioxamine B degrading enzyme is a serine protease. Canadian Journal of Microbiology. 44:521-527, 1998.
Castignetti, D. 1997. Probing of Pseudomonas aeruginosa, Pseudomonas aureofaciens, Burkholderia (Pseudomonas) cepacia, Pseudomonas fluorescens and Pseudomonas putida with the ferripyochelin receptor A gene and the synthesis of pyochelin in Pseudomonas aureofaciens, Pseudomonas fluorescens and Pseudomonas putida. Curr. Microbiol. 34:250-257.
Harwani, S.C.,A. Roginsky, Y. Vallejo and D. Castignetti. 1997. Further characterization and proposed pathway of deferrioxamine B catabolism. BioMetals. 10:205-213.
Fig. Non-denaturing gel of fractions from Sephadex C-25 column. Lanes 1and 4 were inactive fractions (no DFB hydrolase activity) while lanes 2 and 3 were active DFB hydrolase fractions. Note the absence of the top band in lanes 1 and 4 versus lanes 2 and 3. The top band represents DFB hyrdrolase, estimated to be about 110,000 daltons based on the non-denaturing (lanes 5-8) PAGE standards. Size of the standards for the non-denaturing lanes are 14,200, 29,000, 45,000, 66,000, 132,000, 272,000 and 545,000 (alpha-lactalbumin, carbonic anhydrase, chicken egg albumin, bovine serum albumin [monomer at 66K and dimer at 132K], jack bean urease [272K and 545K]). A number of these bands, however, are difficult to see as they are either in the dye front or run as charged isomers. The heavy band in lanes 5-8 is bovine serum albumin monomer (66 kDa) while the next band above is its the dimer, 132 kDa. In lane 5, the band below the bovine serum albumin 66 kDa band is the 45,000 kDa chicken egg albumin and the band below it is the 29 kDa carbonic anhydrase. Analysis of the standards and the unknown bands indicates that the higher molecular weight band in lanes 2 and 3 (DFB hyrdolase) has a molecular weigth of about 110 kDa. The contaminant band has a molecular weight of about 66 kDa.