Methods and kits using a molecular interaction between a Smurf-1 WW domain and LIM mineralization protein isoforms
AssigneeEmory University
InventorsScott D. Boden, Sreedhara Sangadala
Life of this patent
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Abstract
The instant application provides kits and methods for identifying agents which induce or inhibit the osteogenic effect of LMP or BMP proteins. The kits are directed to methods which measure either an amount of a complex between a Smurf protein or a fragment thereof and an LMP protein or a fragment thereof. Alternatively, the kits are directed to methods of measuring an amount of the ubiquitinated Smad protein or a fragment thereof.
Claims51
A method of identifying an agent affecting a binding between an LMP protein and a Smurf1 protein comprising:
- providing a first composition comprising:
- a first amino acid sequence comprising an amino acid sequence of SEQ. ID NO. 1;
- a second amino acid sequence comprising an amino acid sequence at least 70% identical to SEQ. ID. NO. 2 and capable of binding the amino acid sequence of SEQ ID NO 1;
- and
- the agent;
- and
- measuring an amount of a complex formed between the first amino acid sequence and the second amino acid sequence in the first composition.
A method of identifying an agent affecting ubiquitination of a Smad protein by Smurf1 comprising:
- providing a first composition comprising:
- a first amino acid sequence comprising an amino acid sequence of SEQ. ID. NO. 1;
- a second amino acid sequence comprising an amino acid sequence at least 70% identical to SEQ. ID. NO. 2 (WW domain of Smurf1), said second amino acid sequence capable of binding the amino acid sequence of SEQ. ID. NO. 1 and of ubiquitinating the Smad protein;
- the Smad protein;
- a source of ubiquitin;
- a source of ATP;
- the agent;
- and
- measuring an amount of a ubiquitinated Smad protein in the first composition.
A method of identifying an agent affecting ubiquitination of a Smad protein by a Smurf1 protein comprising:
- providing a first composition comprising:
- the Smurf1 protein or a fragment thereof capable of ubiquitinating the Smad protein;
- a source of ubiquitin;
- a source of ATP;
- the Smad protein or a fragment thereof capable of being ubiquitinated by the Smurf1 protein;
- the agent;
- and
- measuring an amount of ubiquitinated Smad protein or the fragment thereof in the first composition.
A kit comprising:
- a first amino acid sequence comprising an amino acid sequence of SEQ. ID. NO. 1;
- and
- a second amino acid sequence comprising an amino acid sequence at least 70% identical to SEQ. ID. NO. 2 (WW domain of SMURF1) and capable of binding the amino acid sequence of SEQ. ID. NO. 1.
A kit comprising
- a Smurf1 protein or a fragment thereof capable of ubiquitinating a Smad protein;
- a source of ubiquitin;
- a source of ATP;
- the Smad protein or a fragment thereof capable of being ubiquitinated by the Smurf1 protein.
A method of identifying an agent affecting a binding between an LMP protein and a Smurf1 protein comprising:
- a) obtaining coordinates for a three-dimensional structure of a Smurf1 protein or a WW-2 motif containing fragment thereof;
- b) selecting the agent by performing a rational drug design with the three-dimensional coordinates, wherein said selecting is performed in conjunction with computer modeling of a complex between an LMP protein or a fragment thereof and the Smurf1 protein or the fragment thereof.
Dependencies are inferred from the claim text.