US 12,392,783 B2
Methods for rapidly digesting biopolymers with ultrastable enzymes for mass spectrometry-based analyses
Steven M. Yannone, Berkeley, CA (US); Jill O. Fuss, Berkeley, CA (US); and Adam Barnebey, Berkeley, CA (US)
Assigned to CINDER BIOLOGICAL, INC., Berkeley, CA (US)
Appl. No. 16/758,806
Filed by CINDER BIOLOGICAL, INC., Berkeley, CA (US)
PCT Filed Oct. 24, 2018, PCT No. PCT/US2018/057397
§ 371(c)(1), (2) Date Apr. 23, 2020,
PCT Pub. No. WO2019/084196, PCT Pub. Date May 2, 2019.
Claims priority of provisional application 62/576,374, filed on Oct. 24, 2017.
Prior Publication US 2021/0063408 A1, Mar. 4, 2021
Int. Cl. C12N 9/52 (2006.01); G01N 33/68 (2006.01)
CPC G01N 33/6842 (2013.01) [C12N 9/52 (2013.01); G01N 33/6848 (2013.01)] 19 Claims
 
1. A method of preparing a biological sample, comprising:
(a) providing the biological sample comprising at least one biopolymer;
(b) contacting the sample with a composition comprising an ultrastable enzyme from an organism of the Archaea domain, to form a reaction mixture, wherein the ultrastable enzyme cleaves the biopolymer at one or more specific sites, and wherein the ultrastable enzyme is selected from the group consisting of SEQ ID NO: 26 and SEQ ID NO: 35; and
(c) incubating the reaction mixture for at least one second to digest the at least one biopolymer present in the biological sample, at a pH between 0.5-7.0 and a temperature between 50° C.-150° C., and
wherein steps (a) to (c) produce a prepared sample for proteomic, glycomic, or glycoproteomic analysis, and wherein the prepared sample is injected into an analytical device for proteomic, glycomic, or glycoproteomic analysis after step (c).