TY - JOUR
T1 - In vivo liquid–liquid phase separation protects amyloidogenic and aggregation-prone peptides during overexpression in Escherichia coli
AU - Gabryelczyk, Bartosz
AU - Alag, Reema
AU - Philips, Margaret
AU - Low, Kimberly
AU - Venkatraman, Anandalakshmi
AU - Kannaian, Bhuvaneswari
AU - Shi, Xiangyan
AU - Linder, Markus
AU - Pervushin, Konstantin
AU - Miserez, Ali
N1 - Publisher Copyright:
© 2022 The Protein Society.
PY - 2022/5
Y1 - 2022/5
N2 - Studying pathogenic effects of amyloids requires homogeneous amyloidogenic peptide samples. Recombinant production of these peptides is challenging due to their susceptibility to aggregation and chemical modifications. Thus, chemical synthesis is primarily used to produce amyloidogenic peptides suitable for high-resolution structural studies. Here, we exploited the shielded environment of protein condensates formed via liquid–liquid phase separation (LLPS) as a protective mechanism against premature aggregation. We designed a fusion protein tag undergoing LLPS in Escherichia coli and linked it to highly amyloidogenic peptides, including β amyloids. We find that the fusion proteins form membraneless organelles during overexpression and remain fluidic-like. We also developed a facile purification method of functional Aβ peptides free of chromatography steps. The strategy exploiting LLPS can be applied to other amyloidogenic, hydrophobic, and repetitive peptides that are otherwise difficult to produce.
AB - Studying pathogenic effects of amyloids requires homogeneous amyloidogenic peptide samples. Recombinant production of these peptides is challenging due to their susceptibility to aggregation and chemical modifications. Thus, chemical synthesis is primarily used to produce amyloidogenic peptides suitable for high-resolution structural studies. Here, we exploited the shielded environment of protein condensates formed via liquid–liquid phase separation (LLPS) as a protective mechanism against premature aggregation. We designed a fusion protein tag undergoing LLPS in Escherichia coli and linked it to highly amyloidogenic peptides, including β amyloids. We find that the fusion proteins form membraneless organelles during overexpression and remain fluidic-like. We also developed a facile purification method of functional Aβ peptides free of chromatography steps. The strategy exploiting LLPS can be applied to other amyloidogenic, hydrophobic, and repetitive peptides that are otherwise difficult to produce.
KW - amyloids
KW - E. coli
KW - liquid–liquid phase separation
KW - membraneless organelles
KW - protein condensates
KW - protein tag
KW - recombinant expression
UR - http://www.scopus.com/inward/record.url?scp=85129220614&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85129220614&partnerID=8YFLogxK
U2 - 10.1002/pro.4292
DO - 10.1002/pro.4292
M3 - Article
C2 - 35481658
AN - SCOPUS:85129220614
SN - 0961-8368
VL - 31
JO - Protein Science
JF - Protein Science
IS - 5
M1 - e4292
ER -