# US 2023/0057500 A1 · Automated lipid exchange-mass spectrometry systems and methods

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- Number: US 2023/0057500 A1
- Document ID: US20230057500A1
- Status: Published application
- Kind: A1
- Application number: 17/796,054
- Filed: 2021-02-04
- Published: 2023-02-23
- Earliest claimed: 2020-02-06
- Assignees: Arizona Board of Regents on Behalf of the University of Arizona
- Applicants: Arizona Board of Regents On Behalf of the University of Arizona
- Inventors: Michael Marty; James Keener; Guozhi Zhang
- Web page: https://patentgrep.com/patent/US20230057500A1

## CPC

- G01N 30/7233: {interfaced to liquid or supercritical fluid chromatograph (interfaces in general for introducing or extracting samples to be analysed with specially adapted mass spectrometer, see H01J49/04)}
- H01J 49/0027: {Methods for using particle spectrometers}
- G01N 30/30: of temperature
- G01N 33/58: involving labelled substances (G01N33/53 takes precedence)
- G01N 33/6848: {Methods of protein analysis involving mass spectrometry}
- G01N 33/92: involving lipids, e.g. cholesterol {, lipoproteins, or their receptors (steroid hormones G01N33/743)}
- G01N 35/0099: {comprising robots or similar manipulators (robots per se B25J)}
- G01N 2030/027: {Liquid chromatography}
- G01N 2333/665: Assays involving proteins derived from pro-opiomelanocortin, pro-enkephalin or pro-dynorphin
- G01N 2500/00: Screening for compounds of potential therapeutic value
- G01N 2560/00: Chemical aspects of mass spectrometric analysis of biological material
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- H01J: ELECTRIC DISCHARGE TUBES OR DISCHARGE LAMPS (spark-gaps H01T; electric arc lamps H05B31/00; particle accelerators H05H)

## Abstract

An automated system for lipid exchange-mass spectrometry, e.g., measuring affinity of a membrane protein for lipids. The automated systems herein can measure the specificity of membrane protein-lipid interactions, detect remodeling of the membrane environment, and determine optimal lipid composition for membrane proteins.

## Claims

### Claim 1

A method for measuring affinity of a membrane protein for lipids, said method comprising:
a) incubating untagged lipid nanoparticles with a population of tagged lipid nanoparticles, wherein the untagged lipid nanoparticles comprise one or more lipids and a membrane protein, wherein the membrane protein is embedded in the untagged lipid nanoparticles, wherein the tagged lipid nanoparticles comprise one or more lipids without the membrane protein embedded therein;
b) separating the tagged lipid nanoparticles from the untagged lipid nanoparticles;
c) measuring a lipid composition of the untagged lipid nanoparticles; and
d) calculating a difference between the lipid composition of the untagged lipid nanoparticles and a lipid composition of the combination of both tagged lipid nanoparticles and untagged lipid nanoparticles;
wherein one or more of steps (a)-(d) are automated.

### Claim 2 (depends on 1)

The method of claim 1, wherein the lipid nanoparticle is a nanodisc.

### Claim 3 (depends on 1)

The method of claim 1, wherein the untagged lipid nanoparticles and the tagged lipid nanoparticles have one or more types of lipids, and wherein the untagged lipid nanoparticles have at least one type of lipid that is different from the one or more types of lipids of the tagged lipid nanoparticles.

### Claim 4

(canceled)

### Claim 5 (depends on 1)

The method of claim 1, wherein step (a) is performed in a temperature-controlled reaction chamber.

### Claim 6 (depends on 1)

The method of claim 1, wherein the method further comprises a robotic mechanism for mixing the untagged lipid nanoparticles with the tagged lipid nanoparticles before incubating the untagged lipid nanoparticles and the tagged lipid nanoparticles.

### Claim 7 (depends on 1)

The method of claim 1, wherein separating the tagged lipid nanoparticles from the untagged lipid nanoparticles comprises a robotic mechanism introducing the mixture of untagged lipid nanoparticles and tagged lipid nanoparticles to a separation system.

### Claim 8

(canceled)

### Claim 9 (depends on 7)

The method of claim 7, wherein the separation system is a chromatography system, wherein the chromatography system is affinity chromatography.

### Claim 10

(canceled)

### Claim 11

(canceled)

### Claim 12 (depends on 7)

The method of claim 7, wherein the separation system removes the tagged lipid nanoparticles and captures the untagged lipid nanoparticles.

### Claim 13 (depends on 12)

The method of claim 12, wherein capturing the untagged lipid nanoparticles comprises a robotic mechanism introducing the untagged lipid nanoparticles to a collection system.

### Claim 14 (depends on 13)

The method of claim 13, wherein the collection system is a chromatography system, wherein the chromatography system is liquid chromatography (LC).

### Claim 15

(canceled)

### Claim 16

(canceled)

### Claim 17 (depends on 13)

The method of claim 13, wherein the untagged lipid nanoparticles captured by the collection system are subsequently subjected to mass spectrometry for measuring the lipid composition of the untagged lipid nanoparticles.

### Claim 18

(canceled)

### Claim 19

(canceled)

### Claim 20 (depends on 1)

The method of claim 1, wherein the method is for determining an optimal lipid environment for a membrane protein.

### Claim 21 (depends on 20)

The method of claim 20, wherein the membrane protein is a protein that is the subject of a drug screening or drug discovery process

### Claim 22

A system (100) for measuring affinity of a membrane protein for lipids, said system comprising:
a) a temperature controlled-reaction chamber (101) for incubating a mixture of tagged lipid nanoparticles and untagged lipid nanoparticles at a selected temperature;
b) a separation system (102) for separating tagged lipid nanoparticles from untagged lipid nanoparticles;
c) a mass spectrometer (103); and
d) one or more robotic mechanisms for introducing lipid nanoparticles to or transferring lipid nanoparticles from one or a combination of (a)-(c).

### Claim 23 (depends on 22)

The system of claim 22, wherein the lipid nanoparticles are nanodiscs.

### Claim 24 (depends on 22)

The system of claim 22, wherein the separation system is a chromatography system, wherein the chromatography system is affinity chromatography.

### Claim 25

(canceled)

### Claim 26

(canceled)

### Claim 27 (depends on 22)

The system of claim 22, wherein the one or more robotic mechanisms are capable of performing one or a combination of the following actions:
a) mixing the mixture in the temperature controlled-reaction chamber;
b) introducing a mixture to the temperature controlled-reaction chamber;
c) transferring the mixture in the temperature controlled-reaction chamber to the separation system; or
d) transferring untagged lipid nanoparticles to the mass spectrometer.

### Claim 28 (depends on 22)

The system of claim 22, wherein the one or more robotic mechanisms are capable of performing one or a combination of the following actions:
a) incubating the untagged lipid nanoparticles having the membrane protein embedded therein with the tagged lipid nanoparticles without the membrane protein embedded therein;
b) separating the tagged lipid nanoparticles from the untagged lipid nanoparticles; and
c) measuring a lipid composition of the untagged lipid nanoparticles.

### Claim 29 (depends on 28)

The system of claim 28, wherein the system further comprising a memory having computer executable instructions for calculating a difference between the lipid composition of the untagged lipid nanoparticles and a lipid composition of the combination of both tagged lipid nanoparticles and untagged lipid nanoparticles.

### Claim 30 (depends on 28)

The system of claim 28, wherein the untagged lipid nanoparticles and the tagged lipid nanoparticles have one or more types of lipids, and wherein the untagged lipid nanoparticles have at least one type of lipid that is different from the one or more types of lipids of the tagged lipid nanoparticles.

### Claim 31

(canceled)