Journal
JOURNAL OF BIOTECHNOLOGY
Volume 344, Issue -, Pages 11-23Publisher
ELSEVIER
DOI: 10.1016/j.jbiotec.2021.12.006
Keywords
EcaA; Carbonic anhydrase; Crocosphaera subtropica ATCC 51142; Signal peptide; Heterologous expression; Protein secretion
Categories
Funding
- Russian Foundation for Basic Research, Russia [19-04-00457]
- Ministry of Science and Higher Education of the Russian Federation, Russia [121041200194-7, 121033000137-1]
- Russian Science Foundation, Russia [21-74-30003]
- Russian Science Foundation [21-74-30003] Funding Source: Russian Science Foundation
Ask authors/readers for more resources
This study expressed different forms of EcaA protein in Escherichia coli and found that the recombinant proteins with a leader peptide could form mature EcaA in the outer layers of the cells, showing high extracellular carbonic anhydrase activity. The cyanobacterial signal peptide played an important role in the secretion machinery of E. coli.
Several forms of EcaA protein, correspondent to the extracellular alpha-class carbonic anhydrase (CA) of cyano-bacterium Crocosphaera subtropica ATCC 51142 were expressed in Escherichia coli. The recombinant proteins with no leader peptide (EcaA and its fusion with thioredoxin or glutathione S-transferase) were allocated inside cells in a full-length form; these cells did not display any extracellular CA activity. Soluble proteins (including that of periplasmic space) of E. coli cells that expressed both EcaA equipped with its native leader peptide (L-EcaA) as well as L-EcaA fused with thioredoxin or glutathione S-transferase at N-terminus, mainly contained the processed EcaA. The appearance of mature EcaA in outer layers of E. coli cells expressed leader peptide-containing forms of recombinant proteins, has been directly confirmed by immunofluorescent microscopy. Those cells also displayed high extracellular CA activity. In addition, the mature EcaA protein was detected in the culture medium. This suggests that cyanobacterial signal peptide is recognized by the secretory machinery and by the leader peptidase of E. coli even as a part of a fusion protein. The efficiency of EcaA leader peptide was comparable to that of PelB and TorA signal peptides, commonly used for biotechnological production of extracellular recombinant proteins in E. coli.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available