Methods for identifying agents which alter histone protein acetylation
AssigneeMassachusetts Institute of Technology
InventorsLeonard Guarente, Shin-ichiro Imai, Christopher Armstrong
Life of this patent
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Abstract
Methods of identifying agents which alter the NAD-dependent acetylation status and mono-ADP-ribosylation of nuclear proteins are disclosed. The methods further include identifying agents which alter the life span or aging of a cell or an organism by determining the level of NAD-dependent acetylation and/or ADP ribosylation of a nuclear protein. The invention also relates to a mammalian Sir2 protein which acetylates or deacetylates nuclear proteins in a NAD-dependent manner and has mono-ADP-ribosyltransferase activity. Host cells producing the Sir2 protein and antibodies to the Sir2 protein are also provided.
Claims49
A method of evaluating the deacetylation of a histone protein in the presence of:
- (i) a Sir2 protein, (ii) NAD or an NAD-like compound, and (iii) an agent, the method comprising:
- a) combining:
- (i) a histone protein that comprises an acetylated amino acid side chain,
- (ii) an isolated or recombinantly produced Sir2 protein,
- (iii) NAD or an NAD-like compound, and
- (iv) an agent to be tested,
- thereby producing a combination;
- and
- b) determining if the histone protein is deacetylated.
A method of evaluating the deacetylation of a histone protein or an N-terminal fragment of an H3 or an H4 histone protein in the presence of a human Sir2 protein, NAD, and an agent, the method comprising:
- a) providing a mixture comprising
- (i) a histone protein or an N-terminal fragment of an H3 or an H4 histone protein that comprises an acetylated amino acid side chain, wherein the N-terminal fragment comprises amino acids 1-20 of SEQ ID NO:6;
- SEQ ID NO:7;
- or SEQ ID NO:8,
- (ii) an isolated or recombinantly produced human Sir2 protein,
- (iii) NAD, and
- (iv) an agent to be tested;
- and
- b) determining if the histone protein or the N-terminal fragment of an H3 or an H4 histone protein is deacetylated.
A method of evaluating the deacetylation of an N-terminal fragment of an H3 or an H4 histone protein in the presence of:
- (i) a Sir2 protein, (ii) NAD or an NAD-like compound, and (iii) an agent, the method comprising:
- a) combining:
- (i) an N-terminal tail of an H3 or an H4 histone protein that comprises an acetylated amino acid side chain, wherein the N-terminal tail comprises amino acids 1-20 of SEQ ID NO:6;
- SEQ ID NO:7;
- or SEQ ID NO:8,
- (ii) an isolated or recombinantly produced Sir2 protein,
- (iii) NAD or an NAD-like compound, and
- (iv) an agent to be tested,
- thereby producing a combination;
- and
- b) determining if the N-terminal tail of an H3 or H4 histone protein is deacetylated.
Dependencies are inferred from the claim text.