Isobaric tandem mass tags for quantitative proteomics and peptidomics

Patent metadata

Field	Value
Publication number	US-10119974-B2
Application number	US-201615182329-A
Country	US
Kind code	B2
Filing date	Jun 14, 2016
Priority date	Sep 15, 2011
Publication date	Nov 6, 2018
Grant date	Nov 6, 2018

Abstract

Compositions and methods of tagging peptides and other molecules using novel isobaric tandem mass tagging reagents, including novel N, N-dimethylated amino acid 8-plex and 16-plex isobaric tandem mass tagging reagents. The tagging reagents comprise: a) a reporter group having at least one atom that is optionally isotopically labeled; b) a balancing group, also having at least one atom that is optionally isotopically labeled, and c) an amine reactive group. The tagging reagents disclosed herein serve as attractive alternatives for isobaric tag for relative and absolute quantitation (iTRAQ) and tandem mass tags (TMTs) due to their synthetic simplicity, labeling efficiency and improved fragmentation efficiency.

First claim

The invention claimed is:

1. A kit comprising two or more tagging reagents, wherein each tagging reagent comprises a reporter group, an amine reactive group, and a balancing group located between the reporter group and amine reactive group, wherein one or more atoms in the reporter group, balancing group, or both, are isotopically heavy versions of the atom, wherein, for each isotopically heavy version of the atom, heavy isotopes are present in excess of the naturally occurring isotopic abundance;
wherein the reporter group of each tagging reagent has a mass different than the reporter groups of the other tagging reagents, the balancing group of each tagging reagent has a mass different than the balancing groups of other tagging reagents, and the aggregate mass of the reporter group plus the balancing group for each tagging reagent is the same;
wherein a first tagging reagent has the formula:
(CH 3 ) 2 -AA 1 -AA 2 -R 1   (formula 16)

where R 1 is an amine reactive group;
AA 1 is a first amino acid having an N-terminus;
AA 2 is a second amino acid having an N-terminus wherein AA 1 and AA 2 cannot be the same amino acid; and
the two CH 3 groups are attached to the N-terminus of AA 1 ; and
wherein at least one tagging reagent has the formula:
(CH 3 ) 2 -AA 2 -AA 1 -R 1   (formula 17)

where AA 1 and AA 2 are the same amino acids as in the first tagging reagent with the exception that the amino acids may contain different isotopes; and
the two CH 3 groups are attached to the N-terminus of AA 2 ;

wherein the reporter group of each tagging reagent has the formula:

wherein R 2 and R 3 , independently of one another, are CH 3 , 13 CH 3 , CDH 2 , 13 CDH 2 , CD 2 H, 13 CD 2 H, CD 3 or 13 CD 3 ;

wherein the balancing group of each tagging reagent has the formula:

wherein R 4 and R 5 are, independently from one another, selected from the group consisting of hydrogen, deuterium, a methyl group optionally containing one or more deuterium atoms and a 13 C atom; an isopropyl group optionally containing one or more deuterium atoms and one or more 13 C atoms; and a butyl group optionally containing one or more deuterium atoms and one or more 13 C atoms;
R 6 is hydrogen or deuterium;
C V and C x , independently of one another, are 12 C or 13 C,
O U and O y , independently of one another, are 16 O or 18 O;
N W is 14 N or 15 N; and

wherein at least one of: R 2 or R 3 contains a deuterium atom, N z is 15 N, or N W is 15 N; and at least one of O U or O y is 18 O.

2. The kit of claim 1 wherein AA 1 and AA 2 , independently from one another, are selected from the group consisting of leucine, isoleucine, alanine, glycine, and valine with the provision that AA 1 and AA 2 cannot have the same mass.

3. The kit of claim 1 wherein the reporter group of each tagging reagent has a mass that differs from one another by two or more Daltons.

4. The kit of claim 1 comprising from 2 to 8 tagging reagents having the formula (CH 3 ) 2 -AA 1 -AA 2 -R 1 and from 2 to 8 tagging reagents having the formula (CH 3 ) 2 -AA 2 -AA 1 -R 1 .

5. The kit of claim 1 wherein the reporter group of each tagging reagent has a mass that differs from one another by one or more Daltons.

Assignees

• Wisconsin Alumni Res Found

Inventors

• Li Lingjun
• Xiang Feng
• Greer Tyler
• Frost Dustin
• Liang Zhidan

Classifications

• C07D207/46 Primary

  with hetero atoms directly attached to the ring nitrogen atom · CPC title

• G01N33/6848 Primary

  Methods of protein analysis involving mass spectrometry · CPC title