Description of Data Sets


MIPS General The Munich Information Center For Protein Sequences is a curated, comprehensive database of yeast proteins. In addition to providing functional and sequence data, the database also provides the user with multiple protein-protein interaction tables. Additionally, the MIPS database catalogues Yeast two-hybrid interactions. MIPS
Physical & Genetic These include: (i) Genetic Interactions e.g. synthetic lethality- where only the knockout of a combination of two genes, but not each individual gene, results in a lethal phenotype indicating functional association. And (ii), physical Interactions e.g. co-purification - where a protein is extracted with another protein indicating that both proteins physically interact in vivo. Physical

Yeast two-hybrid Interactions Limited Ito et al and Uetz et al produced genome-wide libraries of yeast two hybrid interactions, first established by O. Song and S. Fields. In this method for determining protein interactions, two sets of fusion proteins (one to a DNA binding domain and the other to a transcriptional activation domain) are allowed to interact. In a case where the two proteins do interact, the combined DNA binding domain and the transcriptional activation domains will transcribe a reporter gene, usually GAL4. The strength of the Y2H technique lies in its ability to detect both weak and or transient interactions in a true in vivo setting. The method is known to produce many false positives, and all interactions must take place in the nucleus. Ito et al

Uetz et al
Genome-wide The genome wide yeast two hybrid libraries differ from the more limited directed experiments in that they are not conducted by fishing for ‘bait’ proteins with a single ‘prey’ protein, but rather they use the entire genome as prey to fish through the baited genome. As such these libraries are limited by the annotation of the genome and may include random fragments, and may not be fully comprehensive.
Biochemical Studies Chemical Cross Linking Chemical cross-linking of RNA polymerase II subunits was performed with five different cross-linkers, dimethylsuberimidate (DMS), 2-iminothiolane hydochloride (ILT), N, N'- o -phenylene dimaleimide (PDM), dimethyl 3,3'-dithio-bis propionimidate (DTBP) and diexpoxybutane (DEB). Crosslinking subuniots were then detected on a gel by Western blotting with antibodies to RNA Pol II subunits. Ishiguro et al
Far Western In these experiments RNA polymerase II subunits were resolved by denaturing gel electrophoresis, transferred to a solid support, renatured and over-laid with various purified subunits
Pull-Down In these experiments every recombinant subunit was cloned into baculoviruses as either a GST-tagged or an untagged protein. Every combination of individual GST-tagged subunits and non-tagged subunits were co-expressed and precipitated, generating a comprehensive set of inter-subunit pairwise interactions Acker et al
Kimura & Ishihama
Larkin & Guilfoyle
Ulmasov et al
Yasui et al
In-vivo Pull Down TAP Tagging Gavin et al use a Tandem-Affinity Purification, methodology to isolate yeast protein complexes for subsequent mass spectrometry analysis. This method allows for the elucidation of many of the component proteins in a complex without any prior knowledge of the complex composition, activity, or function. The group, using 1,739 tagged genes identified 98 known nonredundant, multiprotein complexes and 134 new complexes ranging from 2 to 83 components.TAP tagging may not always find a complex if it is not formed during the specific cell cycle stage that is being tested, or under the given conditions. Gavin et al
HMS PCI High-throughput mass spectrometric protein complex identification, carried out by Ho et al with 725 bait proteins, determined 1,578 interacting proteins in yeast. This method, which is similar to the TAP tagging method, found many protein complexes, was determined to be more efficient and powerful than conventional yeast two hybrid experiments. The method is limited in that weakly interacting proteins may not survive the purification step. Ho et al