Modification of protein by use of a controllable interveining protein sequence
AssigneeNew England Biolabs, Inc.
InventorsDonald G. Comb, Francine B. Perler, William E. Jack, Ming-Qun Xu, Robert A. Hodges
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Abstract
The present invention is directed to modified proteins and methods of their production. The modified proteins comprise a controllable intervening protein sequence (CIVPS) inserted into a target protein, the CIVPS being capable of excision from the modified protein under predetermined conditions, i.e., increase in temperature, exposure to light, unblocking of amino acid residues by dephosphorylation or deglycosylation. If desired, the modified protein can be subjected to these conditions. The CIVPS may also be inserted into a region that substantially inactivates target protein activity.
Claims36
A modified protein comprising a target protein into which is inserted a controllable intervening protein sequence, wherein said controllable intervening protein sequence is capable of excision under predetermined conditions, and wherein said modified protein is non-naturally occurring.
A method of producing a modified protein which comprises:
- (a) inserting a DNA encoding a controllable intervening protein sequence into a DNA encoding a target protein to form a fusion DNA;
- (b) expressing said fusion DNA to produce the modified target protein,
A method of producing a protein comprising:
- (a) inserting a DNA encoding a controllable intervening protein sequence into a DNA encoding a target protein, wherein said controllable intervening protein sequence is capable of excision under predetermined conditions;
- (b) expressing the DNA of step (a) to produce a modified target protein;
- and
- (c) subjecting the modified target protein to conditions under which said controllable intervening protein sequence will undergo excision.
A method of producing a protein comprising:
- (a) producing a first modified protein comprising an amino portion of a target protein into which is inserted at its carboxy terminus a controllable intervening protein sequence;
- (b) producing a second modified protein comprising the remaining portion of the target protein of step (a) into which is inserted at its amino terminus a controllable intervening protein sequence;
- and
- (c) placing the first and second modified proteins under predetermined conditions appropriate for splicing of the controllable intervening protein sequence.
A method of producing a protein comprising:
- (a) producing a first modified protein comprising a portion of a target protein into which a controllable intervening protein sequence is inserted at a splice junction;
- and
- (b) placing the first modified protein with the remaining portion of the target protein under predetermined conditions appropriate for splicing of the controllable intervening protein sequence.
Dependencies are inferred from the claim text.