Multiple effects of an RNA polymerase β' mutation on in vitro transcription

Abstract

S. tyrphimurium strain BY324 is temperature sensitive due to a mutation (rpo C32) in the gene for the RNA polymerase beta' subunit. Transcription of T7 DNA by RNA polymerase purified from this strain is temperature sensitive in vitro. The enzyme is slightly defective in template binding and RNA chain initiation, but the major defect is in RNA chain elongation. The rate of RNA chain elongation is reduced 4-5 fold relative to wild-type. RNA chain termination does not appear to be affected by the beta' mutation. While the elongation defect is suppressed by glycerol or dimethylsulfoxide, the initiation defect is not. Possible roles for the beta' subunit in enzyme function are discussed in light of these results.