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Shapiro, Stuart Z. Alm, Elizabeth Wheeler 2011-05-07T12:37:07Z 2011-05-07T12:37:07Z 10000-01-01 1992 Phosphorylation of proteins at tyrosine is an important mechanism for regulating cell growth and proliferation in metazoan organisms. This phosphorylation is conducted by a tyrosine-specific protein kinase. The activation of tyrosine kinases is an important step in transducing extracellular signals to a cell's interior. Until recently, protozoan organisms were thought not to possess tyrosine-specific protein kinases. This investigation demonstrates that Trypanosoma brucei, a protozoan parasite, has a tyrosine-specific protein kinase that plays a role in regulation of proliferation of this protozoan. Genistein, a tyrosine kinase inhibitor, prevented multiplication of the parasite and caused the trypanosomes to accumulate in the G2 or M phase of the cell cycle. An in vitro kinase assay demonstrated the presence of a kinase capable of phosphorylating an exogenous substrate at tyrosine, and genistein was able to reduce trypanosome-mediated phosphorylation of this substrate. Two-dimensional, thin-layer electrophoresis of alkali-hydrolyzed ($\sp{32}$P) -labeled total trypanosome protein revealed the presence of phosphotyrosine. An immunoblot of trypanosome proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and incubated with an anti-phosphotyrosine monoclonal antibody revealed the presence of a tyrosine phosphoprotein. An alkali digestion of ($\sp{32}$P) -labeled trypanosome proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated several additional proteins that may be phosphorylated at tyrosine. The phosphorylation of some of these proteins was sensitive to the inhibitor genistein. These results indicate that T. brucei has a tyrosine kinase that is involved in proliferation or growth regulation of the parasite and provide further evidence for the possibility of growth factor regulation and signal transduction in trypanosomes. Made available in DSpace on 2011-05-07T12:37:07Z (GMT). No. of bitstreams: 2 license.txt: 4922 bytes, checksum: 910b249b4beec47e7ab768910c8f966f (MD5) 9215765.pdf: 3833393 bytes, checksum: a3d0781e776de07db0ca8c5005665ba0 (MD5) Previous issue date: 1992 Item marked as restricted to the 'UIUC Users [automated]' Group (id=2) by Howard Ding (hding2@illinois.edu) on 2011-05-07T14:43:24Z Item is restricted indefinitely. Restriction data tranferred 2014-07-01T11:18:59-05:00 Original Data Group with Access UIUC Users [automated] Release Date: none Reason: ETDs are only available to UIUC Users without author permission ETDs are only available to UIUC Users without author permission U of I Only AAI9215765 (UMI)AAI9215765 http://hdl.handle.net/2142/20365 eng Copyright 1992 Alm, Elizabeth Wheeler Biology, Cell Agriculture, Animal Pathology Biology, Veterinary Science Growth regulation in the protozoan parasite Trypanosoma brucei: Evidence of tyrosine-specific protein kinase activity text Pathobiology Pathobiology University of Illinois at Urbana-Champaign Dissertation Ph.D.