ISOLATION OF THE GENE ENCODING PYROCOCCUS-FURIOSUS ORNITHINE CARBAMOYLTRANSFERASE AND STUDY OF ITS EXPRESSION PROFILE IN-VIVO AND IN-VITRO

The gene coding for ornithine carbamoyltransferase (OTCase, argF) in t he hyperthermophilic archaea Pyrococcus furiosus was cloned by complem entation of an OTCase mutant of Escherichia coli. The cloned P. furios us argF gene also complemented a similar mutant of Saccharomyces cerev isiae. Sequencing revealed an open reading frame of 314 amino acids ho mologous to known OTCases and preceded by a TATA box showing only limi ted similarity with the Euryarchaeota consensus sequence. This is in a ccordance with the comparatively low in vitro promoter activity observ ed in a cell-free purified transcription system. Transcription initiat es in vivo as well as in vitro at a guanine, 22 nucleotides downstream of the TATA box. Upstream from argF is a putative gene for diphthine synthetase, a eukaryotic enzyme assumed to occur also in archaea but n ot in bacteria.