Low-salt crystallization of T7 RNA polymerase: a first step towards the transcription bubble complex.
DNA-dependent RNA polymerase is the key enzyme responsible for the biosynthesis of RNA, a process known as transcription. This process, which decodes the genetic information from DNA, is one of the most significant events in a biological system. The crystallization of both native and a chimeric T7/T3 RNAP using high salt conditions has been reported previously but these conditions proved unsuitable for DNA-RNAP complex formation since at high salt concentrations the DNA binding affinity to RNAP is reduced. A search for low-salt crystallization conditions has yielded new low-salt crystals of native T7-RNAP, a chimeric T7-RNAP (T7/T3 RNAP) which contains the T3 promoter recognition sequence, and a T7-RNAP containing an N-terminal histidine tag. The crystals, which are better suited for DNA-RNAP complex formation, belong to space group P3121 with a = 136, c = 156 A, contain a single molecule per asymmetric unit and diffract to 2.7 A resolution. Packing analysis shows that the new low-salt crystals have packing contacts similar to those observed in the high-salt T7-RNAP crystals reported previously. The diffraction anisotropicity observed in crystals of T7 RNAP is explained in term of crystal packing.
D001483:Base Sequence; D003460:Crystallization; D017931:DNA Primers; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry*; D006639:Histidine / Q000737:chemistry; D008958:Models, Molecular; D011487:Protein Conformation; D014158:Transcription, Genetic; D014764:Viral Proteins