Characterization and transcriptional regulation of the 2'-N-acetyltransferase gene from Providencia stuartii.
We have cloned the chromosomally encoded 2'-N-acetyltransferase gene [aac(2')-Ia] from Providencia stuartii. DNA sequence analysis of the cloned insert identified a single open reading frame, which is capable of encoding a protein with a predicted molecular mass of 20,073 Da. The deduced AAC(2')-Ia protein showed no significant homology to other proteins, including all of the AAC(3) and AAC(6') proteins. Primer extension analysis was used to identify the aac(2')-Ia promoter, which contained an unusual sequence (CTTTTT) at the -35 region. Expression of the aac(2')-Ia gene occurs at low levels in wild-type P. stuartii strains; therefore, they are aminoglycoside susceptible. We have isolated mutants with high-level AAC(2')-Ia expression at a frequency of 4.8 x 10(-6). Detailed analysis of one mutant demonstrated a 12.2-fold increase in the accumulation of aac(2')-Ia mRNA. In addition, the levels of beta-galactosidase expression from a plasmid-encoded aac(2')-lacZ transcriptional fusion were increased 11.5-fold in this mutant relative to those in an isogenic wild-type strain. These results suggested that a trans-acting factor, designated aar (for aminoglycoside acetyltransferase regulator), controls AAC(2')-Ia expression in P. stuartii.
Rather
P N
PN
Department of Medicine, Case Western Reserve University School of Medicine, Cleveland, Ohio.
Orosz
E
E
Shaw
K J
KJ
Hare
R
R
Miller
G
G
D000123:Acetyltransferases / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002876:Chromosomes, Bacterial / Q000648:ultrastructure; D004352:Drug Resistance, Microbial; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005809:Genes, Regulator; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D011401:Promoter Regions, Genetic; D011532:Providencia / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D014158:Transcription, Genetic
D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D000123:Acetyltransferases; C443113:aminoglycoside 2'-N-acetyltransferase