| 摘要 |
A method for recombinantly expressing a macromolecule in a host cell is disclosed which involves culturing a host cell which contains two nucleic acid sequences, i.e., a first nucleic acid sequence encoding a membrane-permeabilizing agent and a second nucleic acid sequence encoding a desired macromolecule under the operative control of an inducible promoter, to a selected cell density that permits accumulation of the agent. Thereafter the host cell is exposed to an environmental condition that induces the agent to disrupt the integrity of the cell membrane without complete lysis of the cell membrane. The host cell thereby allows transport through the membrane of small molecular weight compounds. These resulting host cells are cultured in the presence of a nutrient cocktail that contains components that can transport through the disrupted cell membrane, e.g., an inducing agent that induces the tightly regulated promoter and metabolic requirements that permit expression of the macromolecule. Alternatively, a method for enhancing recombinant expression of a macromolecule in a host cell comprises contacting a host cell at a suitable cell density with a membrane-permeabilizing agent that disrupts the integrity of the cell membrane without complete lysis of the membrane, and allows transport through the membrane of small molecular weight compounds. The host cell contains a nucleic acid sequence encoding a macromolecule under the operative control of an inducible promoter. These cells are then cultured in the presence of an above-described nutrient cocktail, and permits enhanced expression of the macromolecule in the membrane-disrupted host cell. Each method can also be employed in methods for in situ drug screening, among other uses. |
