Abstract
The nucleotide sequence of the Myxococcus xanthus orotidine-5'-monophosphate decarboxylase (OMP DCase) gene was determined. The derived protein sequence is not closely related to other prokaryotic OMP DCase sequences; nor is it closely related to any eukaryotic OMP DCase sequences. Progressive multiple alignment of the M. xanthus OMP DCase protein sequence with 19 other OMP DCase sequences revealed four conserved regions present in all 20 sequences. Ten entirely conserved residues were found in these four regions and one region contains a tight cluster of 5 conserved residues, certain of which may be catalytically active residues. A second open reading frame was found upstream of uraA and oriented in the same direction as uraA. A stretch of 21 consecutive pyrimidine (C or T) residues were found in the intercistronic region between the potential ribosome-binding site of uraA and the UGA stop codon of the upstream open reading frame. RNA directly upstream of the pyrimidine run, including the UGA stop codon of the upstream open reading frame, could be folded into a stable hairpin structure resembling Rho-independent terminators of Escherichia coli. Expression of the uraA gene may be regulated by an intercistronic transcription termination mechanism.
Highlights
The nucleotide sequence of thMeyxococcus xanthus in diverse fungi and in the slime mold Dictyosteleum discoiorotidine-5’-monophosphate decarboxylase
Ten entirely conserved residues were found in these four regionsand one region contains a tight clustoefr 6 conserved residues, certain of whimchay becatalytwith the derived protein sequence of the M. xanthus enzyme, all 20 known OMP DCase sequences were aligned using new computer-aided sequence alignment methods
The Escherichia coli pyrFmutant strain open reading frame, coulbde folded into a stable hair- BNN46 was used for complementation testing of cloned M. xanthus pin structure resembling Rho-independent terminatorDsNA fragments
Summary
Transformed into E. coli host strain XLI-blue (Stratagene,Inc.). The enzyme orotidine-5’-monophosphatedecarboxylase (OMP DCase)’ catalyzes the final step of de noua pyrimidine nucleotide biosynthesis [1].The decarboxylation catalyzed by OMP DCase is unusual in that itprobably does not proceed reactions were performed using modified T7 DNA polymerase [5]and [%S]dATP (1,300Ci/mmol), according to theprotocols provided with the Sequenase 2.0 sequencing kit (United States Biochemicals). Reaction products were fractionated on 0.4-mm thick 6 or 8%polyacrylamide sequencing gels [6].Regions where the sequence was obscured via a pyridoxal phosphate:enzyme covalent intermediate, as by strong band compression occurred on average once per template. Except for that of Bacillus subtilis,were obtained from the GENBANK, NBRF or SWISSPROT sequence databases. The nucleotidesequence(s) reported in this papehras been submitted to the GenBankTM/EMBLData Bank withaccessionnumber(s)
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