Genes and enzymes of ectoine biosynthesis in halotolerant methanotrophs.

Ectoine (1,4,5,6-tetrahydro-2-methyl-4-pyrimidine carboxylic acid) is a widely distributed compatible solute accumulated by halophilic and halotolerant microorganisms to prevent osmotic stress in highly saline environments. Ectoine as a highly water keeping compound stabilizing biomolecules and whole cells can be used in scientific work, cosmetics, and medicine. Detailed understanding of the organization/regulation of the ectoine biosynthetic pathway in various producers is an active area of research. Here we review current knowledge on some genetic and enzymatic aspects of ectoine biosynthesis in halophilic and halotolerant methanotrophs. By using PCR methodology, the genes coding for the specific enzymes of ectoine biosynthesis, diaminobutyric acid (DABA) aminotransferase (EctB), DABA acetyltransferase (EctA), and ectoine synthase (EctC), were identified in several methanotrophic species. Organization of these genes in either ectABC or ectABC-ask operons, the latter additionally encoding aspartate kinase isozyme (Ask), correlated well with methanotroph halotolerance and intracellular ectoine level. A new gene, ectR1 encoding the MarR-like transcriptional regulatory protein EctR1, negatively controlling transcription of ectoine biosynthetic genes was found upstream of ectABC-ask operon in Methylomicrobium alcaliphilum 20Z. The ectR-like genes were also found in halotolerant methanol utilizers Methylophaga alcalica and Methylophaga thalassica as well as in several genomes of nonmethylotrophic species. The His(6)-tagged DABA acetyltransferases from Mm. alcaliphilum, M. alcalica, and M. thalassica were purified and the enzyme properties were found to correlate with the ecophysiologies of these bacteria. All these discoveries should be very helpful for better understanding the biosynthetic mechanism of this important natural compound, and for the targeted metabolic engineering of its producers.