Affiliation:
1. Department of Microbiology, University of Iowa, Iowa City, Iowa 52242
Abstract
ABSTRACT
Three transposon Tn
5367
mutagenesis vectors (phAE94, pPR28, and pPR29) were used to create a collection of insertion mutants of
Mycobacterium bovis
strain BCG. A strategy to select for transposon-generated mutants that cannot make coenzyme F
420
was developed using the nitroimidazopyran-based antituberculosis drug PA-824. One-third of 134 PA-824-resistant mutants were defective in F
420
accumulation. Two mutants that could not make F
420
-5,6 but which made the biosynthesis intermediate FO were examined more closely. These mutants contained transposons inserted in two adjacent homologues of
Mycobacterium tuberculosis
genes, which we have named
fbiA
and
fbiB
for F
420
biosynthesis. Homologues of
fbiA
were found in all seven microorganisms that have been fully sequenced and annotated and that are known to make F
420
.
fbiB
homologues were found in all but one such organism. Complementation of the
fbiA
mutant with
fbiAB
and complementation of the
fbiB
mutant with
fbiB
both restored the F
420
-5,6 phenotype. Complementation of the
fbiA
mutant with
fbiA
or
fbiB
alone did not restore the F
420
-5,6 phenotype, but the
fbiA
mutant complemented with
fbiA
produced F
420
-2,3,4 at levels similar to F
420
-5,6 made by the wild-type strain, but produced much less F
420
-5. These data demonstrate that both genes are essential for normal F
420
-5,6 production and suggest that the
fbiA
mutation has a partial polar effect on
fbiB
. Reverse transcription-PCR data demonstrated that
fbiA
and
fbiB
constitute an operon. However, very low levels of
fbiB
mRNA are produced by the
fbiA
mutant, suggesting that a low-level alternative start site is located upstream of
fbiB
. The specific reactions catalyzed by FbiA and FbiB are unknown, but both function between FO and F
420
-5,6, since FO is made by both mutants.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Cited by
84 articles.
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