A gene encoding an extracellular lipase from Streptomyces sp. M11 was cloned in the high-copy-number vector pIJ486, using S. lividans 66 as host. A 28-kDa protein was secreted by S. lividans carrying pB13, which harbors a 6-kb insert, and identified as the product of the cloned gene. Comparison of the N-terminal amino acid (aa) sequence of the purified extracellular lipase with the nucleotide (nt) sequence of the lip gene revealed the presence of a 48 aa long signal peptide. The nucleotide sequence also revealed the presence of a motif, Gly-His-Ser-Met-Gly, similar to the one found surrounding the active-site Ser in other lipases. The gene is most likely monocistronic. Subcloning experiments indicated that another gene might be required for high-level expression, since subcloning of the structural gene alone resulted in diminished extracellular lipase activity. The lipase gene promoter was identified by S1 mapping experiments, and found to be similar to other Streptomyces vegetative promoters.
1. Starvation for 3 days causes membrane damage of the rat erythrocyte manifested by several alterations. The adenosine-triphosphatase activity is decreased but that of acetylcholinesterase is not affected. 2. The ouabain-sensitive adenosine-triphosphatase activity increases at the expense of the non-sensitive enzyme moiety. 3. The Rb(+) uptake is not altered but the galactose transport is accelerated by the stated experimental conditions. 4. The modifications induced by starvation do not recover on re-feeding.
