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Method for engineering proteases and protein kinases

  • US 9,546,359 B2
  • Filed: 12/19/2014
  • Issued: 01/17/2017
  • Est. Priority Date: 06/25/2012
  • Status: Active Grant
First Claim
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1. A nucleic acid vector for engineering protease variants, wherein the nucleic acid vector encodes two separately expressed proteins, wherein the first protein is a first fusion protein comprising, in an N- to C-terminal direction:

  • (i) a first endoplasmic reticulum (ER) targeting sequence;

    (ii) a surface expression sequence;

    (iii) a first peptide sequence that is a counterselection substrate sequence for the enzyme of the second protein;

    (iv) a first epitope tag sequence;

    (v) a second peptide sequence that is a selection substrate sequence for the enzyme of the second fusion protein;

    (vi) a second epitope tag sequence; and

    (vii) a first endoplasmic reticulum (ER) retention sequence;

    wherein the second protein is a protease or has protease activity.

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