Summary: Genes for lactose catabolism and proteinase production in 712 are encoded by a 56.5 kb metabolic plasmid, pLP712. A lactose mini-plasmid of only 23.7 kb, pMG820, was constructed by introducing two deletions into pLP712, and was cloned as two segments of DNA into the vector pAT153 using restriction endonuclease I. The lactose genetic region of pLP712, which has been defined by deletion and restriction mapping, was cut into two parts by this process. When the smaller 10.8 kb segment of pMG820 DNA was present, the key lactic streptococcal lactose splitting enzyme, phospho-β-d-galactosidase, was expressed in The gene for phospho-β-d-galactosidase was more precisly located by introducing a series of deletions into cloned DNA by manipulations and then assaying for enzyme activity. The presence of this phospho-β-d-galactosidase activity was correlated with the production of a 58kDa S-labelled protein both by minicells and after coupled transcription and translation of cloned DNA. The product of a second gene, a 37 kDa protein (“protein X”), and a possible truncated phospho-β-d-galactosidase protein of 16 kDa were also detected in minicells.


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