High-efficient bacterial production of human ApoA-I amyloidogenic variants
(2018) In Protein Science 27(12). p.2101-2109- Abstract
Apolipoprotein A-I (ApoA-I)-related amyloidosis is a rare disease caused by missense mutations in the APOA1 gene. These mutations lead to protein aggregation and abnormal accumulation of ApoA-I amyloid fibrils in heart, liver, kidneys, skin, nerves, ovaries, or testes. Consequently, the carriers are at risk of single- or multi-organ failure and of need of organ transplantation. Understanding the basic molecular structure and function of ApoA-I amyloidogenic variants, as well as their biological effects, is, therefore, of great interest. However, the intrinsic low stability of this type of proteins makes their overexpression and purification difficult. To overcome this barrier, we here describe an optimized production and purification... (More)
Apolipoprotein A-I (ApoA-I)-related amyloidosis is a rare disease caused by missense mutations in the APOA1 gene. These mutations lead to protein aggregation and abnormal accumulation of ApoA-I amyloid fibrils in heart, liver, kidneys, skin, nerves, ovaries, or testes. Consequently, the carriers are at risk of single- or multi-organ failure and of need of organ transplantation. Understanding the basic molecular structure and function of ApoA-I amyloidogenic variants, as well as their biological effects, is, therefore, of great interest. However, the intrinsic low stability of this type of proteins makes their overexpression and purification difficult. To overcome this barrier, we here describe an optimized production and purification procedure for human ApoA-I amyloidogenic proteins that efficiently provides between 46 mg and 91 mg (depending on the protein variant) of pure protein per liter of Escherichia coli culture. Structural integrity of the amyloidogenic and native ApoA-I proteins were verified by circular dichroism spectroscopy and intrinsic fluorescence analysis, and preserved functionality was demonstrated by use of a lipid clearance assay as well as by reconstitution of high-density lipoprotein (HDL) particles. In conclusion, the use of the described high-yield protein production system to obtain amyloidogenic ApoA-I proteins, and their native counterpart, will enable molecular and cellular experimental studies aimed to explain the molecular basis for this rare disease.
(Less)
- author
- Del Giudice, Rita LU and Lagerstedt, Jens O. LU
- organization
- publishing date
- 2018
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- amyloidosis, apolipoprotein A-I, HDL, high-density lipoprotein, high-yield protein production
- in
- Protein Science
- volume
- 27
- issue
- 12
- pages
- 9 pages
- publisher
- The Protein Society
- external identifiers
-
- scopus:85056591086
- pmid:30291643
- ISSN
- 0961-8368
- DOI
- 10.1002/pro.3522
- language
- English
- LU publication?
- yes
- id
- e58737d6-1437-4259-8765-3e420560f354
- date added to LUP
- 2018-11-26 13:25:46
- date last changed
- 2024-04-29 19:24:12
@article{e58737d6-1437-4259-8765-3e420560f354,
abstract = {{<p>Apolipoprotein A-I (ApoA-I)-related amyloidosis is a rare disease caused by missense mutations in the APOA1 gene. These mutations lead to protein aggregation and abnormal accumulation of ApoA-I amyloid fibrils in heart, liver, kidneys, skin, nerves, ovaries, or testes. Consequently, the carriers are at risk of single- or multi-organ failure and of need of organ transplantation. Understanding the basic molecular structure and function of ApoA-I amyloidogenic variants, as well as their biological effects, is, therefore, of great interest. However, the intrinsic low stability of this type of proteins makes their overexpression and purification difficult. To overcome this barrier, we here describe an optimized production and purification procedure for human ApoA-I amyloidogenic proteins that efficiently provides between 46 mg and 91 mg (depending on the protein variant) of pure protein per liter of Escherichia coli culture. Structural integrity of the amyloidogenic and native ApoA-I proteins were verified by circular dichroism spectroscopy and intrinsic fluorescence analysis, and preserved functionality was demonstrated by use of a lipid clearance assay as well as by reconstitution of high-density lipoprotein (HDL) particles. In conclusion, the use of the described high-yield protein production system to obtain amyloidogenic ApoA-I proteins, and their native counterpart, will enable molecular and cellular experimental studies aimed to explain the molecular basis for this rare disease.</p>}},
author = {{Del Giudice, Rita and Lagerstedt, Jens O.}},
issn = {{0961-8368}},
keywords = {{amyloidosis; apolipoprotein A-I; HDL; high-density lipoprotein; high-yield protein production}},
language = {{eng}},
number = {{12}},
pages = {{2101--2109}},
publisher = {{The Protein Society}},
series = {{Protein Science}},
title = {{High-efficient bacterial production of human ApoA-I amyloidogenic variants}},
url = {{http://dx.doi.org/10.1002/pro.3522}},
doi = {{10.1002/pro.3522}},
volume = {{27}},
year = {{2018}},
}