Compounds and methods used in assessing mono-PARP activity

Patent metadata

Field	Value
Publication number	US-11530235-B2
Application number	US-201916715907-A
Country	US
Kind code	B2
Filing date	Dec 16, 2019
Priority date	Nov 20, 2015
Publication date	Dec 20, 2022
Grant date	Dec 20, 2022

Abstract

Mutant mono ADP-ribose-polymerases (mono-PARP) proteins and small molecule compound substrates specific for the mutant mono-PARP proteins as well as methods of using these compositions to identify protein targets of the mono-PARPs and to screen for antagonists of the mono-PARPs are described.

First claim

The invention claimed is:

1. A method of identifying a protein target of a mono-PARP, the method comprising: contacting a small molecule compound (SMC) of formula:

wherein R is alkyl or aryl, with a protein, said protein comprising a polypeptide comprising a mono-poly-ADP-ribose-polymerases (PARP) catalytic domain comprising SEQ ID NO: 15, and wherein the polypeptide catalyzes the addition of the SMC to a PARP protein target, wherein said contacting occurs within a living cell; subjecting the cell to conditions that result in the protein catalyzing the reaction by which the SMC is covalently attached to one or more cellular proteins; and identifying the one or more cellular proteins to which the SMC is covalently attached as the protein target(s) of the mono-PARP.

2. The method of claim 1 , wherein the SMC comprises 5-Bn-6-a-NAD+.

3. The method of claim 1 , further comprising conjugating a label to the SMC.

4. The method of claim 3 , wherein the label comprises biotin or a fluorescent molecule.

5. The method of claim 1 , further comprising detecting the one or more cellular proteins to which the SMC is covalently attached via a method that comprises mass spectrometry.

6. The method of claim 1 , wherein R is aryl.

7. The method of claim 6 , wherein R is benzyl.

8. The method of claim 1 , wherein R is alkyl.

9. The method of claim 8 , wherein R is C 1-6 alkyl.

10. The method of claim 9 , wherein R is selected from the group of methyl, isobutyl, and propyl.

Assignees

• Univ Oregon Health & Science

Inventors

• Cohen Michael
• Carter-O'Connell Ian
• Morgan Rory
• Jin Haihong

Classifications

• C07H19/207 Primary

  the phosphoric or polyphosphoric acids being esterified by a further hydroxylic compound, e.g. flavine adenine dinucleotide or nicotinamide-adenine dinucleotide · CPC title

• C12Q1/48

  involving transferase · CPC title

• G01N2500/10

  involving cells · CPC title

• C12Y204/0203

  NAD+ ADP-ribosyltransferase (2.4.2.30), i.e. tankyrase or poly(ADP-ribose) polymerase · CPC title

• C12N9/1077

  Pentosyltransferases (2.4.2) · CPC title