No. 37, 2003 iF, pp. 19-24 Research Reports of the School of Engineering, Kinki University No. 37,2003, pp. 19-24 J: Q fRl __ RNA !J:J. 7 -it(/) iiiJ£1J Q The effect of hydrostatic pressure on activity of RNA polymerase from piezophile by in vitro transcription Kaoru Nakasone We have compared pressure-induced subunit dissociation of RNA polymerase of piezophilic Shewanella violacea and mesophilic Escherichia coli. The transcriptional activity of the purified pressurized-RNA polymerase from both species was measured by both nonspecific and specific in vitro transcription assay, using Poly (dA-dT)·(dA-dT) and the S. violacea rpoA promoter as a DNA template. These analyses showed the RNA polymerase from S. violacea is stable to pressure- induced inactivation compared with E. coli, suggesting molecular adaptation to high pressure environment (piezosphere). Keywords RNA polymerase, Transcription, High pressure, dissociation In vitro transcription 1. Introduction The piezophilic deep-sea bacterium Shewanella violacea DSSI2, isolated from the Ryukyu trench (depth; 5110 m), grows optimally at 30 MPa and ST, but also at atmospheric pressure (0.1 MPa) and ST [1,2]. It is useful as a model bacterium for comparison of various features of bacterial physiology under high and low pressure conditions. Recently an operon identified as a pressure- regulated operon, with a promoter activated under high pressure conditions, was cloned and characterized from this 19 strain [3-5). We have reported that gene expression from this operon is controlled at the transcriptional level by elevated pressure [3]. This piezophilic bacterium, S. violacea is one of deep-sea adapted organisms, can grow at the range of 0.1 MPa to 70 MPa, suggesting almost of the protein structures in this piezopble are more stable and retain the activity required to lives under high hydrostatic pressure conditions than that in organisms living under atmospheric conditions. Department of Biotechnology and Chemistry, School of Engineering, Kinki University