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Methods for the genetic manipulation of the unicellular, nitrogen-fixing cyanobacteria Gloeothece and Synechocystis are being developed to produce modified strains that can be used in physiological and biochemical studies of the interactions, at the molecular level, between nitrogen fixation and oxygen.
Progress has been made in developing genetic systems that will be of use in biochemical and physiological studies into nitrogen fixation by unicellular cyanobacteria. Two genera have been investigated: Gloeothece, which is capable of aerobic nitrogen fixation and a strain of Synechocystis, which requires very low concentrations of oxygen in order to fix nitrogen. A genomic library of Gloeothece deoxyribonucleic acid (DNA) has now been constructed. Unfortunately, however, transformation of Gloeothece using plasmids that successfully replicate in other cyanobacteria remains difficult, even in a strain that lacks the extensive slime capsule that normally surrounds this cyanobacterium. However, chromosomal transfer has been demonstrated with Gloeothece, and this technique may be more suitable for future genetic work with this organism.
In contrast, a plasmid-based conjugative gene-transfer system for Synechocystis has been described and optimized. Chromosomal transformation has also been demonstrated for this organism. A large number of mutant strains of Gloeothece have been studied at the physiological and biochemical level. Many mutants that are incapable of aerobic nitrogen fixation are impaired in respiratory oxygen consumption, lacking in particular a specific, kinetically identifiable, low-affinity oxidase. This may correlate with a deficiency in certain soluble cytochromes. On the other hand, at least one mutant strain has been identified that cannot fix nitrogen aerobically yet still contains this low-affinity oxidase. Studies with a new class of mutants, resistant to agents that induce oxidative stress, have suggested that the enzymes superoxide dismutase and ascorbate peroxidase also have an important role in limiting oxidative damage in Gloeothece.
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