![]() ![]() ![]() For example, the TFIID complex is involved in pre-initiation complex assembly for RNA polymerase II transcription and is composed of at least 13 subunits in humans. Interestingly, those components of the general transcription machinery that function directly in core promoter DNA binding are often composed of large multi-subunit complexes. The initial step of promoter recognition by the general transcription machinery is a key step in transcription and is frequently targeted for intervention during gene regulation. In eukaryotic organisms, transcription occurs by three different RNA polymerases I, II, and III that are unable to directly recognize promoter elements but instead require distinct assemblies of general transcription factors for efficient promoter recruitment. This co-expression system will facilitate the functional characterization of this unusual multi-protein transcription factor that plays an important early role for transcription by two different polymerases. Importantly, this partial complex functions in reconstituted transcription of both human U1 and U6 snRNA genes by RNA polymerases II and III, respectively. The co-expressed complex binds DNA specifically and recruits TBP to U6 promoter DNA. Herein, we describe the co-expression in Escherichia coli of a partial SNAP C containing SNAP190 (1–505), SNAP50, SNAP43, and SNAP19. Expression in a baculovirus system yields active complex but the concentration of such material is insufficient for many bio-analytical methods. Previous studies have shown that a partial SNAP C composed of SNAP190 (1–514), SNAP50, and SNAP43 expressed in baculovirus is capable of PSE-specific DNA binding and transcription of human snRNA genes by RNA polymerases II and III. ![]() SNAP C consists of five subunits: SNAP190, SNAP50, SNAP45, SNAP43, and SNAP19. This multi-subunit complex recognizes the proximal sequence element (PSE) commonly found in the upstream promoters of human snRNA genes. ![]() Regardless of polymerase specificity, these genes share a requirement for a general transcription factor called the snRNA activating protein complex or SNAP C. Human small nuclear (sn) RNA genes are transcribed by either RNA polymerase II or III depending upon the arrangement of their core promoter elements. ![]()
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