Compositions and methods for targeted delivery to cells
US-2024390271-A1 · Nov 28, 2024 · US
Single-domain antibodies with improved thermal stability under cytoplasmic expression
US10294283B2 · US · B2
| Field | Value |
|---|---|
| Publication number | US-10294283-B2 |
| Application number | US-201414158208-A |
| Country | US |
| Kind code | B2 |
| Filing date | Jan 17, 2014 |
| Priority date | Jan 17, 2014 |
| Publication date | May 21, 2019 |
| Grant date | May 21, 2019 |
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Abstract
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Methods of producing a single-domain antibody (sdAb) include causing a bacteria to express the sdAb into cytoplasm of the bacteria, wherein the sdAb is expressed as a fusion protein with the acid tail of α-synuclein; and then purifying the sdAb, wherein the fusion protein is expressed free of a periplasmic location tag. Such antibodies have the unexpected ability to refold after thermal denaturation.
First claim
Opening claim text (preview).
What is claimed is: 1. A method of producing a single-domain antibody (sdAb), the method comprising: causing a bacteria to express the sdAb into cytoplasm of the bacteria, wherein the sdAb is expressed as a fusion protein with the acid tail of α-synuclein; and then purifying the sdAb, wherein the fusion protein is expressed free of a periplasmic location tag. 2. The method of claim 1 , wherein the sdAb lacks a disulfide bond. 3. The method of claim 1 , further comprising subjecting the sdAb to thermal denaturation and subsequently cooling the sdAb whereby the sdAb substantially refolds into its functional conformation. 4. The method of claim 1 , wherein the fusion protein further comprises an affinity tag for purification. 5. The method of claim 4 , further comprising removing the affinity tag from the sdAb prior to purifying the sdAb. 6. The method of claim 4 , wherein the affinity tag is polyhistidine. 7. The method of claim 1 , wherein the acid tail of α-synuclein comprises the amino acid sequence DPDNEAYEMPSEEGYQDYEPEA (SEQ ID No: 1). 8. The method of claim 1 , further comprising removing the acid tail of α-synuclein from the sdAb prior to purifying the sdAb.
Assignees
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Classifications
Micrococcaceae (F); Staphylococcaceae (F), e.g. Staphylococcus (G) · CPC title
Single domain, e.g. dAb, sdAb, VHH, VNAR or nanobody® · CPC title
Bacillus (G) · CPC title
against material from plants · CPC title
Stability, e.g. half-life, pH, temperature or enzyme-resistance · CPC title
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- What does patent US10294283B2 cover?
- Methods of producing a single-domain antibody (sdAb) include causing a bacteria to express the sdAb into cytoplasm of the bacteria, wherein the sdAb is expressed as a fusion protein with the acid tail of α-synuclein; and then purifying the sdAb, wherein the fusion protein is expressed free of a periplasmic location tag. Such antibodies have the unexpected ability to refold after thermal denatur…
- Who is the assignee on this patent?
- Zabetakis Daniel, Anderson George P, Goldman Ellen R, and 3 more
- What technology area does this patent fall under?
- Primary CPC classification C07K14/47. Mapped technology areas include Chemistry & Metallurgy.
- When was this patent published?
- Publication date Tue May 21 2019 00:00:00 GMT+0000 (Coordinated Universal Time) (B2). Legal status and post-grant events are not shown on this page.
- What related patents are in patentsdb?
- We list 8 related publications on this page (citations in our corpus or others sharing the same primary CPC).