Enterococcus faecalis V583 contains a cytochrome bd-type respiratory oxidase

Abstract

We have cloned an Enterococcus faecalis gene cluster, cydABCD, which when expressed in Bacillus subtilis results in a functional cytochrome bd terminal oxidase. Our results indicate that E. faecalis V583 cells have the capacity of aerobic respiration when grown in the presence of heme.

FIG. 1

Light absorption difference (dithionite-reduced minus air-oxidized) spectra of E. faecalis membranes (3 mg of protein per ml). (A) Membranes from cells grown in the presence of 8 μM hemin; (B) membranes from cells grown in the absence of added hemin. The vertical bar indicates the absorption scale. Difference spectra obtained after oxidation by potassium ferricyanide were identical to those obtained by air oxidation.

FIG. 2

Map of plasmid pLUF04, carrying the E. faecalis cydABCD genes. Pcyd indicates the B. subtilis cyd promoter region. BamHI/BglII shows where the PCR fragment (cut with BglII) was ligated to pCYD26 (cut with BamHI). The chloramphenicol and erythromycin resistance genes are indicated by cat and ermC, respectively.

FIG. 3

Light absorption difference (dithionite-reduced minus ferricyanide-oxidized) spectra of B. subtilis membranes (2.5 mg of protein per ml). (A) LUW20/pCYD26; (B) LUW20/pLUF04 (E. faecalis cydABCD); (C) LUW20/pCYD23 (B. subtilis cydABCD). The vertical bar indicates the absorption scale.

FIG. 4

Light absorption difference (dithionite-reduced minus ferricyanide-oxidized) spectra of B. subtilis membranes (2.5 mg of protein per ml). (A) LUW174 (ΔqoxABCD::kan ΔcydABCD::tet pLUF04), (B) LUW173 (ΔqoxABCD::kan ΔcydABCD::tet pCYD23). The vertical bar indicates the absorption scale.