# US 8,569,455 B2 · Cloning, yeast expression, purification and biological activity of the extension region of the soybean 7S globulin alfa' subunit involved in HEP G2 cell cholesterol homeostasis

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- Number: US 8,569,455 B2
- Document ID: US8569455B2
- Status: Granted
- Kind: B2
- Application number: 13/378,605
- Filed: 2010-06-16
- Published: 2012-07-05
- Granted: 2013-10-29
- Earliest claimed: 2009-06-17
- Assignees: Indena S.p.A.
- Applicants: Paolo Morazzoni; Antonella Riva; Cesare Ponzone; Davide Berlanda; Marcello Duranti; Alessandro Consonni
- Inventors: Paolo Morazzoni; Antonella Riva; Cesare Ponzone; Davide Berlanda; Marcello Duranti; Alessandro Consonni
- Web page: https://patentgrep.com/patent/US8569455B2

## CPC

- C07K 14/415: from plants
- A61P 3/06: Antihyperlipidemics
- A61P 9/10: for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS

## Abstract

A truncated form of α′ chain (eα′), the soybean 7S globulin, active in controlling the cholesterol and triglyceride homeostasis in in vitro and in vivo models, was cloned and expressed in the yeast Pichia pastoris. The recombinant polypeptide spanned 142 amino acid residues from the N-terminal side and included the N-terminal extension region of the soybean alpha′ subunit. The eα′ polypeptide was purified by conventional biochemical techniques and its potential to modulate the activity of the LDL-receptor was evaluated in a human hepatoma cell line (Hep G2) by monitoring the uptake and degradation of labeled LDL.

## Claims

### Claim 1

A recombinant N-terminal hydrophilic fragment of Soybean 7S Globulin α′ Subunit, wherein said recombinant N-terminal hydrophilic fragment of Soybean 7S Globulin α′ Subunit consists of the amino acid sequence of SEQ ID NO: 1.

### Claim 2 (depends on 1)

A process for preparing the N-terminal hydrophilic fragment of Soybean 7S Globulin α′ Subunit of claim 1, comprising:
expressing a polynucleotide encoding the N-terminal hydrophilic fragment of Soybean 7S Globulin α′ Subunit of claim 1 in Pichia pastoris, and isolating the recombinant N-terminal hydrophilic fragment.

### Claim 3 (depends on 1)

A composition comprising as the active ingredient the N-terminal hydrophilic fragment of the Soybean 7S Globulin α′ Subunit of claim 1 and a carrier.