Novel tetracycline resistance determinant from the oral metagenome

Abstract

A major drawback of most studies on how bacteria become resistant to antibiotics is that they concentrate mainly on bacteria that can be cultivated in the laboratory. In the present study, we cloned part of the oral metagenome and isolated a novel tetracycline resistance gene, tet(37), which inactivates tetracycline.

FIG. 1.

Multiple sequence alignment of the Tet 37 sequence with other oxidoreductases by using CLUSTAL W (version 1.82). Flavoprotein from F nucleatum subsp. nucleatum ATCC 255 (GenBank accession number AE010563.1) had 44% identity; Rubredoxin-oxygen oxidoreductase from Desulfovibrio gigas (GenBank accession number Q9F0J6) had 29% identity. Amino acid sequence positions are indicated with numbers, and the blocks named A, B, and C show the possible ADP binding sites, metal binding, and the redox center flavodoxin-like domain (I-YGTM-GNTE----S), respectively. The conserved arginine residue is underlined in the sequence, and the conserved active-site residues are marked with an asterisk.

FIG. 2.

Comparison of the visible absorption UV-Vis spectrum. Spectral changes were observed upon adding NADPH (1 mM) to incubation mixtures containing tetracycline (30 μg/ml) and cell extract of E. coli containing tet(X) before incubation (-×-) and after 90 min of incubation (-○-) and containing tet(37) before incubation (—) and after 90 min of incubation (--▵--). E. coli cell extracts containing tetracycline and the vector only showed no change in absorbance after 90 min of incubation (—▪—). O.D., optical density.