Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Recombinant dna technique included in method of making a...
Patent
1991-07-11
1993-06-29
Schwartz, Richard A.
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Recombinant dna technique included in method of making a...
435 694, 435 6952, 435 696, 435 697, 4351723, 435189, 435195, 435215, 435216, 435226, C12P 2102, C12N 1562, C12N 1558, C12N 1553
Patent
active
052234088
ABSTRACT:
A screening method for the selection of mutagenized proteins that are normally secreted by cells is described. The method includes the development of a cloning vector for the expression of secretory proteins as fusion proteins on the cell surface of transfected mammalian cells. The secreted protein is displayed on the cell surface by fusion with the glycophospholipid membrane anchor of decay accelerating factor (DAF). Tissue-type plasminogen activator (t-PA), which is normally secreted, is used as a model protein. PCR mutagenesis is used to generate random mutations within the Kringle 1 (K1) domain of t-PA. Fluorescence activated cell sorting (FACS) is employed to screen for t-PA mutants possessing a loss of an epitope to a specific Mab, whose nonlinear binding domains overlap with the t-PA clearance receptor contact regions novel t-PA mutants designated N115S, N1425S, and K159R were discovered by this method.
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Goeddel David V.
Leung David W. H.
Rice Glenn C.
Carter Philip W.
Genentech Inc.
Schwartz Richard A.
Winter Daryl B.
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