Affiliation:
1. Department of Biology1and
2. Department of Industrial and Mechanic Engineering,2 University of Rome Three, Rome, Italy, and
3. Division of Industrial Microbiology, Department of Food Science, Wageningen Agricultural University, Wageningen, The Netherlands3
Abstract
ABSTRACT
The M1 strain, able to grow on β-myrcene as the sole carbon and energy source, was isolated by an enrichment culture and identified as a
Pseudomonas
sp. One β-myrcene-negative mutant, called N22, obtained by transposon mutagenesis, accumulated (
E
)-2-methyl-6-methylen-2,7-octadien-1-ol (or myrcen-8-ol) as a unique β-myrcene biotransformation product. This compound was identified by gas chromatography-mass spectrometry. We cloned and sequenced the DNA regions flanking the transposon and used these fragments to identify the M1 genomic library clones containing the wild-type copy of the interrupted gene. One of the selected cosmids, containing a 22-kb genomic insert, was able to complement the N22 mutant for growth on β-myrcene. A 5,370-bp-long sequence spanning the region interrupted by the transposon in the mutant was determined. We identified four open reading frames, named
myrA
,
myrB
,
myrC
, and
myrD
, which can potentially code for an aldehyde dehydrogenase, an alcohol dehydrogenase, an acyl-coenzyme A (CoA) synthetase, and an enoyl-CoA hydratase, respectively.
myrA
,
myrB
, and
myrC
are likely organized in an operon, since they are separated by only 19 and 36 nucleotides (nt), respectively, and no promoter-like sequences have been found in these regions. The
myrD
gene starts 224 nt upstream of
myrA
and is divergently transcribed. The
myrB
sequence was found to be completely identical to the one flanking the transposon in the mutant. Therefore, we could ascertain that the transposon had been inserted inside the
myrB
gene, in complete agreement with the accumulation of (
E
)-2-methyl-6-methylen-2,7-octadien-1-ol by the mutant. Based on sequence and biotransformation data, we propose a pathway for β-myrcene catabolism in
Pseudomonas
sp. strain M1.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology
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