Reconstitution of 30S ribosomal subunits in vitro using ribosome biogenesis factors.

Reconstitution of ribosomes in vitro from individual ribosomal proteins provides a powerful tool for understanding the ribosome assembly process including the sequential incorporation of ribosomal proteins. However, conventional assembly methods require high-salt conditions for efficient ribosome assembly. In this study, we reconstituted 30S ribosomal subunits from individually purified ribosomal proteins in the presence of ribosome biogenesis factors.

Photo-regulation of transcription by RNA polymerase with azobenzene-tethered promoter.

Photo-regulation of transcription reaction by T7-RNA polymerase was investigated by introducing an azobenzene residue into the non-template strand of T7-promoter. It was found that transcription was strongly interfered under dark or visible light irradiation, whereas it efficiently proceeded after UV irradiation. With this photo-responsive promoter, on-off switching of transcription by light irradiation was achieved.

Photo-regulation of RNA polymerase reaction by use of modified DNA carrying an azobenzene.

Transcription reaction by T7-RNA polymerase was photo-regulated on the basis of two strategies as depicted in Scheme 1. It was found that incorporation reaction of azobenzene-tethered uridine triphosphate proceeded only when azobenzene took trans-form (Scheme 1(A)). On the other hand, the transcription was more efficient when the azobenzene moiety, tethered to the non-template strand of the promoter DNA, was in its cis-form under UV irradiation (Scheme 1(B)).