In situ lipid synthesis for protein reconstitution

Patent metadata

Field	Value
Publication number	US-11052044-B2
Application number	US-201816489783-A
Country	US
Kind code	B2
Filing date	Mar 1, 2018
Priority date	Mar 2, 2017
Publication date	Jul 6, 2021
Grant date	Jul 6, 2021

Abstract

Described herein, inter alia, are compositions and methods useful for the reconstitution of membrane proteins.

First claim

What is claimed is:

1. A method for forming a lipid membrane comprising a membrane protein, the method comprising:
adding a reactive maltoside surfactant comprising a reactive thioester moiety to a first solution which comprises the membrane protein; thereby forming a second solution which comprises the reactive maltoside surfactant and the membrane protein; and
(ii) adding a cysteine-functionalized phosphatidylcholine compound to the second solution, thereby forming a synthetic phospholipid; wherein the synthetic phospholipid forms a lipid membrane comprising the membrane protein.

2. The method of claim 1 , comprising solubilizing the membrane protein in a first surfactant to form the first solution.

3. The method of claim 2 , wherein the first surfactant is a nonionic alkyl glucoside.

4. The method of claim 2 , wherein the first surfactant is n-dodecyl-β-D-maltoside.

5. The method of claim 1 , wherein the said reactive maltoside surfactant has the formula:

wherein R 1 is a substituted or unsubstituted alkyl.

6. The method of claim 1 , wherein the cysteine-functionalized phosphatidylcholine compound is a cysteine-functionalized oleoyl lysophosphatidylcholine compound or a cysteine-functionalized palmitoyl lysophosphatidylcholine compound.

7. The method of claim 1 , wherein the cysteine-functionalized phosphatidylcholine compound has the formula:

wherein R 2 is a substituted or unsubstituted alkyl.

8. The method of claim 7 , wherein R 2 is an unsubstituted C 4 -C 20 alkyl or an unsubstituted C 6 -C 18 alkenyl.

9. The method of claim 1 , wherein the synthetic phospholipid has the formula:

wherein R 1 is a substituted or unsubstituted alkyl.

10. The method of claim 9 , wherein R 1 is an unsubstituted C 4 -C 20 alkyl or an unsubstituted C 6 -C 18 alkenyl.

11. The method of claim 1 , wherein the membrane protein is a G protein-coupled receptor.

12. The method of claim 1 , wherein the membrane protein is a 5-hydroxytryptamine receptor, an acetylcholine receptor, an adenosine receptor, an angiotensin receptor, an apelin receptor, a bile acid receptor, a bombesin receptor, a bradykinin receptor, a cannabinoid receptor, a chemerin receptor, a chemokine receptor, a cholecystokinin receptor, a Class A Orphan receptor, a dopamine receptor, an endothelin receptor, a formyl peptide receptor, a free fatty acid receptor, a galanin receptor, a ghrelin receptor, a glycoprotein hormone receptor, a gonadotrophin-releasing hormone receptor, a G protein-coupled estrogen receptor, a histamine receptor, a hydroxycarboxylic acid receptor, a kisspeptin receptor, a leukotriene receptor, a lysophospholipid receptor, a lysophospholipid S1P receptor, a melanin-concentrating hormone receptor, a melanocortin receptor, a melatonin receptor, a motilin receptor, a neuromedin U receptor, a neuropeptide FF/neuropeptide AF receptor, a neuropeptide S receptor, a neuropeptide W/neuropeptide B receptor, a neuropeptide Y receptor, a neurotensin receptor, an opioid receptor, an opsin receptor, an orexin receptor, an oxoglutarate receptor, a P2Y receptor, a platelet-activating factor receptor, a prokineticin receptor, a prolactin-releasing peptide receptor, a prostanoid receptor, a proteinase-activated receptor, a QRFP receptor, a relaxin family peptide receptor, a somatostatin receptor, a succinate receptor, a tachykinin receptor, a thyrotropin-releasing hormone receptor, a trace amine receptor, a urotensin receptor, a vasopressin receptor, or a combination of two or more thereof.

13. The method of claim 1 , wherein the lipid membrane is a liposome.

14. The method of claim 1 , wherein the molar ratio of the membrane protein to the synthetic phospholipid is from about 0.1:1 to about 1:600.

15. A liposome produced by the method of claim 1 .

Assignees

• Univ California

Inventors

• Brea Fernandez Roberto Javier
• Cole Christian
• Devaraj Neal K
• Lyda Brent
• Sunahara Roger

Classifications

• C07H15/14

  attached to a sulfur, selenium or tellurium atom of a saccharide radical · CPC title

• A61K9/127

  Synthetic bilayered vehicles, e.g. liposomes or liposomes with cholesterol as the only non-phosphatidyl surfactant · CPC title

• C07F9/106 Primary

  Adducts, complexes, salts of phosphatides · CPC title

• A61K9/1277 Primary

  Preparation processes; Proliposomes · CPC title

• C07K14/705

  Receptors; Cell surface antigens; Cell surface determinants {(tumour specific antigens C07K14/4748)} · CPC title