	US 12,221,648 B2
	Substrates for covalent tethering of proteins to functional groups or solid surfaces
Carolyn W. Hitko, Grover Beach, CA (US); Thomas Kirkland, Atascadero, CA (US); Sergiy Levin, San Luis Obispo, CA (US); Poncho Meisenheimer, San Luis Obispo, CA (US); Rachel Friedman Ohana, Madison, WI (US); Harry Tetsuo Uyeda, Los Osos, CA (US); and Ji Zhu, Croton on Hudson, NY (US)
Assigned to Promega Corporation, Madison, WI (US)
Filed by Promega Corporation, Madison, WI (US)
Filed on Jul. 13, 2021, as Appl. No. 17/374,460.
Application 17/374,460 is a continuation of application No. 14/207,993, filed on Mar. 13, 2014, granted, now 11,072,812.
Claims priority of provisional application 61/788,257, filed on Mar. 15, 2013.
Prior Publication US 2021/0340593 A1, Nov. 4, 2021
This patent is subject to a terminal disclaimer.
Int. Cl. C12Q 1/34 (2006.01); C12N 9/14 (2006.01); G01N 33/543 (2006.01)

CPC C12Q 1/34 (2013.01) [C12N 9/14 (2013.01); G01N 33/54353 (2013.01)]

14 Claims

1. A method to determine the presence or amount of a mutant dehalogenase in a sample, wherein the mutant dehalogenase is capable of stably binding to a haloalkyl substrate, the method comprising:

(a) contacting the sample with a compound of formula R—L¹-M-L²-A—X, wherein R is a functional group selected from an affinity tag, a fluorophore, a chromophore, a cross-linking group, an amino acid, a peptide, a polypeptide, a nucleotide, and a lipid, wherein R is not 1, 4, 7-triazacyclononane-N, N′, N″-triacetic acid (NOTA), M is a first carbamate group, L¹is a first linker portion that comprises a second carbamate group; wherein the second carbamate group is separated from R by 2 or more linearly connected atoms; wherein the second carbamate group is separated from M by 2 or more linearly connected atoms, L²is a second linker portion, A is (CH₂)₆, X is a halogen, L²-A separates M and X by 6-18 linearly connected atoms, and A—X is a substrate for the mutant dehalogenase;

(b) allowing the mutant dehalogenase to bind to the A—X substrate; and