| Summary: | Strain JG3 is a rhizobacterium isolate originated from root of Zea mays cultivated
in Purwokerto, Indonesia, which has the ability to produce lipase but unclear
taxonomy status. Since lipase catalyzes hydrolysis of triglycerides to form
glycerol and fatty acids, it is intriguing to know whether strain JG3 is able to
metabolize glycerol from the lipolysis process. This study aimed to determine
taxonomy status of strain JG3, to isolate genes involved in glycerol metabolism
(glpD dan glpK) and to identify the ability of the strain to metabolize glycerol.
Bacterial identification was carried out using SEM (Scanning Electron
Microscope), BD Phoenix 100 Automated Microbiology System and 16S rRNA
gene analysis. Partial isolation of glpD and glpK was done using optimized colony
PCR with primers designed from conserved region of referred genes of
Proteobacteria and arbitrarily-primed PCR using gene specific primers.
Identification on the ability of strain JG3 to metabolize glycerol was carried out in
vitro using glycerol minimal medium. Identification test results showed that cells
of strain JG3 are coccobacilli and the rhizobacterium was detected as Alcaligenes
faecalis species with 99% BD Phoenix screening confidence. Phylogenetic
analysis based on 16S rRNA sequence (GenBank acc. no. AB914514) confirmed
this result revealing the closest relationship between strain JG3 and A.faecalis and
A.aquatilis with similarity on nucleotide sequence by 96% maximum. Thus,
strain JG3 belongs to genus Alcaligenes namely Alcaligenes sp. JG3. Colony and
arbitrarily-primed PCR resulted in 3 DNA isolates of 427, 248 and 436 bp, with
GenBank acc. no. of AB862285, AB894421 and AB914513, respectively.
Deduced amino acid sequence analysis showed that 427-bp sequence shared 98%
amino acid similarity with NAD-dependent glycerol-3-phosphate dehydrogenase
(G3PDH) of Stenotrophomonas maltophilia K279a (a key enzym in glycerol
metabolism), 248-bp sequence shared 95% amino acid similarity with DSBA
oxidoreductase of Alcaligenes spp., while 436-bp sequence shared 95% amino
acid similarity with benzoate membrane transporter (BenE) of A.faecalis NCIB
8687. In vitro glycerol utilization test result showed that Alcaligenes sp. JG3 was
able to grow aerobically in minimal medium with glycerol as carbon source, but
not anaerobically. As conclusion, Alcaligenes sp. JG3 is able to metabolize
glycerol aerobically, so that it could be potentially used as effective degradator of
oil and fat wastes.
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