Browsing by Subject "Protein-protein interactions."

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Open Access
Clustering protein-protein interactions based on conversed domain similarities
(2004) Ayaz, Aslı
Protein interactions govern most cellular processes, including signal transduction, transcriptional regulation and metabolism. Saccharomyces ceravisae is estimated to have 16,000 protein interactions. Appereantly only a small number of these interactions were formed ab initio (invention), rest of them were formed through gene duplications and exon shuffling (birth). Domains form functional units of a protein and are responsible for most of the interaction births, since they can be recombined and rearranged much more easily compared to innovation. Therefore groups of functionally similar, homologous interactions that evolved through births are expected to have a certain domain signature. Several high throughput techniques can detect interacting protein pairs, resulting in a rapidly growing corpus of protein interactions. Although there are several efforts for computationally integrating this data with literature and other high throughput data such as gene expression, annotation of this corpus is inadaquate for deriving interaction mechanism and outcome. Finding interaction homologies would allow us to annotate an unannotated interaction based on already annotated known interactions, or predict new ones. In this study we propose a probabilistic model for assigning interactions to homologous groups, according to their conserved domain similarities. Based on this model we have developed and implemented an Expectation-Maximization algorithm for finding the most likely grouping of an interaction set. We tested our algorithm with synthetic and real data, and showed that our initial results are very promising. Finally we propose several directions to improve this work
Open Access
Functional identification of RASGEF1 family of exchange factors as activators of RAP2, and as interacting partners of CCDC124
(2009) Yaman, Elif
Coiled coil domain-124 gene is highly conserved among eukaryotes and the human counterpart encodes a protein with no domain similarities with any previously characterized eukaryotic proteins. In this study, we aimed to identify biological functions and interaction partners of human Ccdc-124. A yeast-two-hybrid analysis carried in this study has revealed that Ccdc-124 interacts with RasGEF1B which was predicted to be a member of Ras guanine exchange factors. The highly conserved RasGEF1 family of proteins contain C-terminal CDC25-homology domain (CDC25- HD) and an N-terminal RasGEF-N domain (Ras Exchange Motif, REM), and is of unknown function and specificity. In this thesis, the interaction of Ccdc-124 and RasGEF1 family of proteins was also established with co-immunoprecipitation and GST pull down assays. On the other hand, by using purified RasGEF1A and RasGEF1B proteins, as well as a large number of Ras family of G-proteins, we established that RasGEF1A and RasGEF1B function as very specific exchange factors for Rap2, a member of the Rap subfamily of Ras-like G-proteins. They do not act on Rap1 or other members of the Ras subfamily. On the other hand, Ccdc-124 protein did not change the stimulatory effect of RasGEF1 family of proteins on any of the tested G proteins in vitro. Furthermore, using reciprocal site-directed mutagenesis, we analyzed residues that allow RasGEF1 proteins to discriminate between Rap1 and Rap2, and we were able to identify Phe39 in the switch I region of Rap2 as a specificity residue. Mutation of the corresponding Ser39 in Rap1 changed the specificity and allowed the nucleotide exchange of Rap1(S39F) to be stimulated by RasGEF1B. This study describes for the first time GEFs that are uniquely specific for Rap2 among Rap family of G-proteins.