THE MELA GENE IS ESSENTIAL FOR MELANIN BIOSYNTHESIS IN THE MARINE BACTERIUM SHEWANELLA-COLWELLIANA

The surface-adhering, Gram-negative marine bacterium Shewanella colwelliana synthesizes a red-brown melanin in the late stage of exponential growth in laboratory culture. Previous studies identified a single gene, melA, from S. colwelliana that could impart the ability to produce melanin to an E. coli host. However, these studies did not demonstrate a requirement for metA during melanization in S. colwelliana. In this paper, genetic analyses, using a broad host range conjugation system to generate specific lesions, reveal that melA null mutants fail to synthesize pigment. The wild-type melA gene provided in trans on a low copy number plasmid complemented these null mutations, as well as a spontaneous pigment variant, to wild-type melanin synthesis. Polyclonal antibodies, raised against a MetA-LacZ fusion protein, were used to confirm the presence of the melA gene product in wild-type S. colwelliana and verify its absence in the non-pigmented mutants. In addition, detection of the MelA protein over the course of growth in batch culture revealed a constant steady-state level of MelA protein, suggesting that the timing of melanization and the quantity of melanin synthesized is not controlled at the level of melA expression.