	US 12,188,059 B2
	Proximity-dependent biotinylation and uses thereof
Steven A. Carr, Cambridge, MA (US); Jason Wright, Cambridge, MA (US); and Sam Myers, Cambridge, MA (US)
Assigned to THE BROAD INSTITUTE, INC., Cambridge, MA (US)
Appl. No. 16/333,185
Filed by THE BROAD INSTITUTE, INC., Cambridge, MA (US)
PCT Filed Sep. 13, 2017, PCT No. PCT/US2017/051428 § 371(c)(1), (2) Date Mar. 13, 2019, PCT Pub. No. WO2018/053053, PCT Pub. Date Mar. 22, 2018.
Claims priority of provisional application 62/394,133, filed on Sep. 13, 2016.
Prior Publication US 2019/0241880 A1, Aug. 8, 2019
Int. Cl. C07H 21/04 (2006.01); C12N 9/00 (2006.01); C12N 9/08 (2006.01); C12N 9/22 (2006.01); C12N 15/10 (2006.01); C12N 15/11 (2006.01); C12N 15/63 (2006.01); G16B 15/30 (2019.01); G16B 20/00 (2019.01)

CPC C12N 9/22 (2013.01) [C12N 9/0065 (2013.01); C12N 9/93 (2013.01); C12N 15/102 (2013.01); C12N 15/11 (2013.01); C12N 15/63 (2013.01); C12Y 111/01011 (2013.01); C12Y 603/04015 (2013.01); G16B 15/30 (2019.02); C07K 2319/09 (2013.01); C07K 2319/85 (2013.01); C12N 2310/20 (2017.05); C12N 2740/16043 (2013.01); G16B 20/00 (2019.02)]

26 Claims

1. A fusion protein comprising:

(a) a nucleotide-editing protein, wherein the nucleotide-editing protein is a Streptococcus pyogenes or Staphylococcus aureus catalytically-inactive Cas9 (dCas9) protein, and

(b) a proximity-dependent protein to catalyze ligation of biotin, or an ATP-biotin interaction loop or a biotin pocket thereof, wherein said proximity-dependent protein is a mutated E. coli Bir A, or a biotin-peroxide dependent ascorbate peroxidase (APEX) or a Split Biotin protein ligase separated at amino-acid positions (1-149, 141-320).