Molecular cloning and functional analysis of a novel tetracycline resistance determinant, tet(V), from Mycobacterium smegmatis.
The nucleotide sequence and mechanism of action of a tetracycline resistance gene from Mycobacterium smegmatis were determined. Analysis of a 2.2-kb sequence fragment showed the presence of one open reading frame, designated tet(V), encoding a 419-amino-acid protein (molecular weight, 44,610) with at least 10 transmembrane domains. A database search showed that the gene is homologous to membrane-associated antibiotic efflux pump proteins but not to any known tetracycline efflux pumps. The steady-state accumulation level of tetracycline by M. smegmatis harboring a plasmid carrying the tet(V) gene was about fourfold lower than that of the parental strain. Furthermore, the energy uncoupler carbonyl cyanide m-chlorophenylhydrazone blocked tetracycline efflux in deenergized cells. These results suggest that the tet(V) gene codes for a drug antiporter which uses the proton motive force for the active efflux of tetracycline. By primer-specific amplification the gene appears to be restricted to M. smegmatis and M. fortuitum.
De Rossi
E
E
Department of Genetics and Microbiology, University of Pavia, 27100 Pavia, Italy.
Blokpoel
M C
MC
Cantoni
R
R
Branzoni
M
M
Riccardi
G
G
Young
D B
DB
De Smet
K A
KA
Ciferri
O
O
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009161:Mycobacterium / Q000187:drug effects* / Q000235:genetics; D013752:Tetracycline / Q000493:pharmacokinetics; D013753:Tetracycline Resistance / Q000235:genetics*
D013752:Tetracycline