MOLECULAR-CLONING OF THE METHANOL DEHYDROGENASE STRUCTURAL GENE FROM METHYLOSINUS-TRICHOSPORIUM OB3B

A 4-kb fragment encoding methanol dehydrogenase (MDH) (EC 1.1.99.8) from Methylosinus trichosporium OB3b has been cloned, with lambda gt11, and expressed in Escherichia coli K12. Organisms infected with recombinant phage express a fusion protein of bigger molecular weight than the purified MDH protein from M. trichosporium OB3b. Subcloning of this fragment into pUC18 allowed identification of a recombinant plasmid, pCIT328, which contained a 2.1-kb fragment that expressed a protein that comigrated with purified MDH on polyacrylamide gels and cross-reacted with the antibody, indicative that the full MDH structural protein was encoded. This fragment also hybridized with an appropriate sized fragment from M. trichosporium OB3b total DNA. © 1990 Springer-Verlag New York Inc.