Degree Type

Dissertation

Date of Award

2000

Degree Name

Doctor of Philosophy

Department

Theses & dissertations (Interdisciplinary)

Major

Molecular, Cellular, and Developmental Biology

First Advisor

Janice E. Buss

Abstract

Covalent attachment of isoprenoid lipids to proteins is a post-translational modification that occurs on approximately 2% of all cellular proteins. Proteins that contain a CaaX motif as their final four amino acids are presumed to be targets for the addition of C15 or C20 isoprenoids by farnesyl transferase (FTase) or geranylgeranyl transferase I (GGTase I) respectively. This modification is generally believed to occur stoichiometrically, and irreversibly, in vivo. Thus, if prenylation is to be regulated, this control must occur prior to lipid addition. At the moment, there is little evidence that the enzymatic activity of prenyl transferases can be modulated. Therefore, cellular control over prenylation must occur at the level of protein substrate preference and/or accessibility. Because preventing prenylation of oncogenic proteins such as Ras holds promise as a new area of cancer chemotherapy, there is a great deal of interest in understanding how prenyl transferases recognize their substrates in vivo. In our efforts to better characterize substrate recognition by FTase and GGTase I, we identified a protein termed murine Guanylate Binding Protein-1 (mGBP1) that appeared to naturally limit its own prenylation. Initial studies showed the impairment in mGBP1 modification was more severe for C20 modification, and that non-CaaX amino acids within the C-terminal region of mGBP1 played a role in limiting prenylation. This C20-specific defect appeared to be an intrinsic property of mGBP1, as mutant proteins with C20-type CaaX motifs were poorly prenylated both in vivo and in vitro. However, the C-terminal 18 amino acids of mGBP1 could function as a suitable substrate for prenylation when appended to the K-Ras4B protein, suggesting that the C-terminus required the participation of other mGBP1 structures in order to avoid prenylation. Removal of up to 400 internal amino acids of mGBP1 did not alleviate the block in prenylation, implicating the N-terminal GTP-binding region of mGBP1 as the inhibitory domain. Therefore, mGBP1 may represent a form of previously unrecognized cellular control over prenylation in which an N-terminal nucleotide-binding domain can negatively influence modification of a C-terminal CaaX motif.

DOI

https://doi.org/10.31274/rtd-180813-744

Publisher

Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/

Copyright Owner

John Thomas Stickney

Language

en

Proquest ID

AAI9962850

File Format

application/pdf

File Size

103 pages

Share

COinS