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Alkanes are oxidized in Acinetobacter sp. strain ADP1 through a three-component monooxygenase, composed of alkane hydroxylase, rubredoxin and rubredoxin reductase. The genes rubA and rubB encode rubredoxin and a NAD(P)H-dependent rubredoxin reductase. It is shown that single base pair substitutions in rubA or rubB lead to defects in alkane degradation, indicating that both genes are essential for alkane utilization.

Localized sets of random point mutations generated by polymerase chain reaction amplification can be transferred to the chromosome of Acinetobacter ADP1 strain in an efficient manner using natural transformation. The natural transformation method eliminates the need for cloning of PCR fragments in plasmids since PCR-amplified DNA fragments are internalized by cells and directly introduced into their genomes by homologous recombination. Results show that homologous recombination of the PCR-amplified DNA fragments into a specific chromosomal docking site from which they can be expressed is made possible by nucleotide sequence identity between the flanking regions and corresponding chromosomal segments in a cloned Acinetobacter recipient.

Acinetobacter PobR and PcaU are transcriptional activators which are very similar to each other based on main structure, DNA-binding sites, metabolic modulators and physiological activity. Mutants were isolated in which the pobR function has been deleted by nucleotide substitutions caused by mistakes during polymerase chain reaction amplification of the gene. Findings indicated that these mutations affect the ability of PobR to bind to its DNA target. PobR and PcaU have a linear domain structure similar to that of LysR transcriptional activators.

A method for the efficient random mutagenesis of selected genes in a bacterial chromosome is studied. The process is a combination of PCR replication errors with the uptake of PCR-amplified DNA via natural transformation. Mutations are also transferred directly to the chromosome through homologous recombination thereby eliminating the need to clone PCR fragments. It is shown that the method can by applied to other genes in Acinetobacter and other natural transformation systems.

16S rRNA gene probes were generated to monitor the growth of Bifidobacterium sp. strain LW420 in infant fecal samples. Sequencing and analysis of DNA from single-strain and mixed cultures of bifidobacteria and from heterogeneous feces samples revealed a significant homology of 16S rDNA of LW420 with known 16S rDNA sequences. PCR analyses showed that all fecal samples was positive for Bifidobacterium.

Expression of the lipA gene regulates lipase production in Acinetobacter calcoaceticus at the genetic level. This was gleaned from an investigation of the involvement of a fatty acyl-responsive DNA-binding protein in the regulation of expression of the A. calcoaceticus lipBA operon. It was also found that the level of active extracellular LipA may be influenced at the level of proteolytic stability or instability of the lipase.

Phenotypic analysis of five Acinetobacter calcoaceticus mutants lacking extracellular lipase, which were generated via insertion mutagenesis, revealed four lipolytic enzymes. Biochemical tests classified four mutants as export mutants, while the additional mutant, designated AAC302, exhibited showed mutation 0.84 kbp upstream of lipA, although it displayed the LipA- phenotype. DNA sequencing of the region revealed an open reading frame that encodes a protein required for the production of extracellular lipase. It is suggested that this protein acts as a lipase-specific chaperone.

The factors controlling the expression of growth-phase-dependent genes are investigated. The cloning of an Acinetobacter calcoaceticus BD413 esterase gene grown on N-broth plates provides more information on the positioning of the gene and its activity. A deletion plasmid is constructed to map the gene on a chromosomal DNA fragment.