β-lactamase genes of gram-negative bacteria
Two β-lactamase gene sequences encoded by Ps. aeruginosa RMS 149 plasmid and Rps. capsulata sp 108 were investigated. The genes were located using DNA recombinant techniques and their nucleic acid sequences were determined using the Sanger dideoxy-sequencing technique. The amino acid sequences were identified and compared with other characterised β-lactamases. They are both class A enzymes (Ambler classification). The pseudomonad plasmid encoded enzyme is expressed constitutively, but its gene sequence has an attenuator sequence - reminiscent of inducible bacterial synthetic operons. It also has three putative loop-forming sequences in the middle of the gene. RNA mapping studies indicate that the attenuator is read through from an upstream promoter. There is low level initiation from its own promoter and the transcripts sometimes terminate around the second internal stem-loop. The full message is also made. Thus, it is likely that the pseudomonad gene is normally highly regulated. Its constitutive expression may be as a result of some control mutation. The rhodopseudomonad enzyme is unlike other characterised Gram-negative class β-lactamases because it is inducible. Gene hybridization experiments suggest that it may be chromosomally encoded in strain sp 108 as well as in the Pen^S strain sp 109. β-lactamase active bands were also observed in PenS Rps. capsulata St. Louis and Rps. sphaeroides. If this is the usual state of affairs in photosynthetic bacteria which are not normally subject to the selective pressures of the presence of β-lactam antibiotics by virtue of their aquatic habitat, the sp 108 strain may also be producing the enzyme in large quantities due to some control mutation. It is postulated then, that β-lactamase genes in Gram-negative bacteria may be of two kinds - one that is chromosomal and is highly regulated, and another which has lost the regulation and over-expresses the enzyme. The latter may be representative of the common plasmid-borne β-lactamase genes.