Prenylation is an important posttranslational modification for many proteins whereby attachment of a lipid tail leads to protein localization to cellular membranes. The two enzymes responsible for the majority of prenylation, farnesyltransferase (FTase) and geranylgeranyltransferase type I (GGTase-l), have been proposed to recognize a consensus """"""""CaaX"""""""" box motif at the C-terminus of target proteins. However, recent work has shown that this model incompletely describes the specificity of these enzymes. Furthermore, the interactions between the protein substrates and FTase responsible for specificity have not been delineated. I will investigate these issues using mutagenesis and directed evolution to generate a library of FTase variants with altered substrate specificities and kinetic behavior. Analysis of these mutated FTases will provide a more comprehensive understanding of the features that govern binding and catalysis within FTase and may aid in the identification of novel substrates and design of new FTase inhibitors as therapeutic agents. In addition, this work will provide important information and novel tools for studying protein prenylation in vivo, allowing examination of the myriad biological roles of prenylated proteins. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32GM078894-02
Application #
7279837
Study Section
Special Emphasis Panel (ZRG1-F04A-D (20))
Program Officer
Fabian, Miles
Project Start
2006-08-01
Project End
2009-01-31
Budget Start
2007-08-01
Budget End
2008-07-31
Support Year
2
Fiscal Year
2007
Total Cost
$48,796
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Hougland, James L; Gangopadhyay, Soumyashree A; Fierke, Carol A (2012) Expansion of protein farnesyltransferase specificity using ""tunable"" active site interactions: development of bioengineered prenylation pathways. J Biol Chem 287:38090-100
London, Nir; Lamphear, Corissa L; Hougland, James L et al. (2011) Identification of a novel class of farnesylation targets by structure-based modeling of binding specificity. PLoS Comput Biol 7:e1002170
Hougland, James L; Hicks, Katherine A; Hartman, Heather L et al. (2010) Identification of novel peptide substrates for protein farnesyltransferase reveals two substrate classes with distinct sequence selectivities. J Mol Biol 395:176-90
Krzysiak, Amanda J; Aditya, Animesh V; Hougland, James L et al. (2010) Synthesis and screening of a CaaL peptide library versus FTase reveals a surprising number of substrates. Bioorg Med Chem Lett 20:767-70
Hougland, James L; Lamphear, Corissa L; Scott, Sarah A et al. (2009) Context-dependent substrate recognition by protein farnesyltransferase. Biochemistry 48:1691-701
Hougland, James L; Piccirilli, Joseph A (2009) 2'-amino-modified ribonucleotides as probes for local interactions within RNA. Methods Enzymol 468:107-25