Cyanobacterial leader peptides for protein secretion

Sergeyenko, Tatiana V; Los, Dmitry A.

doi:10.1016/s0378-1097(02)01197-7

Cited by 15 publications

(9 citation statements)

References 27 publications

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“…It was suggested that these proteins were not essential under the steady state growth condition used in the study, although absence of these proteins affected the rate of Chl synthesis and of the assembly of Chl -binding proteins (He and Vermaas 1999). Sll1694 was also identified as a secreted protein that reached the outer surface of the cell (Sergeyenko and Los 2003). The most definitive work to date has involved motility.…”

Section: Differential Gene Expression In Disia Under Normal Growth Comentioning

confidence: 99%

Novel adaptive responses revealed by transcription profiling of a Synechocystis sp. PCC 6803 ΔisiA mutant in the presence and absence of hydrogen peroxide

Singh

Bono

et al. 2005

Photosynth Res

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The isiAB genes have proven to be highly stress-responsive under a variety of environmental conditions, including iron deficiency, high salt and oxidative stress. In order to understand the function of IsiA and its importance in oxidative stress, we constructed a knock out mutant of the isiA gene and compared differential gene expression of the DeltaisiA strain in the presence and absence of H2O2. We used the full genome microarray for the cyanobacterium Synechocystis sp. PCC 6803 as previously described [Postier BL, Wang HL, Singh A, Impson L, Andrews, HL, Klahn J, Li H, Risinger G, Pesta D, Deyholos M, Galbraith DW, Sherman LA and Burnap RL (2003) BMC Genenomics 4: 23-34]. We determined that one of the main differences in DeltaisiA compared to wild-type (in the absence of peroxide) was the induction of a gene cluster (sll1693-sll1696) that encoded genes resembling pilins or general secretory proteins (Gsp). These proteins are targeted to the cytoplasmic membrane and we suggest that they may be involved in the assembly of membrane complexes, including pigment-protein complexes. The DeltaisiA strain was more resistant to H2O2 compared to the wild-type. In the presence of 1.5 mM H2O2 for 30 min, a cluster of genes that includes a peroxiredoxin was induced 7- to 8-fold and we suggest that this peroxide scavenging enzyme is responsible for the increased peroxide resistance of the DeltaisiA strain.

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Section: Differential Gene Expression In Disia Under Normal Growth Comentioning

confidence: 99%

Novel adaptive responses revealed by transcription profiling of a Synechocystis sp. PCC 6803 ΔisiA mutant in the presence and absence of hydrogen peroxide

Singh

Bono

et al. 2005

Photosynth Res

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“…PCC 6803 by biochemical isolation yielded only a low number of hits and contained periplasmic and outer membrane proteins (Sergeyenko and Los, 2000). By analysing individual proteins, it could be demonstrated that PilA1 (Sll1694), PilA10 (Slr2016), HlyA (Sll1951) as well as CccS (Slr1667) and CccP (Slr1668) are secreted to the extracellular space (Sergeyenko and Los, 2003;Sakiyama et al, 2006;Yoshimura et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

Secretome analysis of Anabaena sp. PCC 7120 and the involvement of the TolC‐homologue HgdD in protein secretion

Hahn

Stevanovic

Brouwer

et al. 2014

Environmental Microbiology

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Secretion of proteins is a central strategy of bacteria to influence and respond to their environment. Until now, there has been very few discoveries regarding the cyanobacterial secrotome or the secretion machineries involved. For a mutant of the outer membrane channel TolC-homologue HgdD of Anabaena sp. PCC 7120, a filamentous and heterocyst-forming cyanobacterium, an altered secretome profile was reported. To define the role of HgdD in protein secretion, we have developed a method to isolate extracellular proteins of Anabaena sp. PCC 7120 wild type and an hgdD loss-of-function mutant. We identified 51 proteins of which the majority is predicted to have an extracellular secretion signal, while few seem to be localized in the periplasmic space. Eight proteins were exclusively identified in the secretome of wild-type cells, which coincides with the distribution of type I secretion signal. We selected three candidates and generated hemagglutinin-tagged fusion proteins which could be exclusively detected in the extracellular protein fraction. However, these proteins are not secreted in the hgdD-mutant background, where they are rapidly degraded. This confirms a direct function of HgdD in protein secretion and points to the existence of a quality control mechanism at least for proteins secreted in an HgdD-dependent pathway.

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“…After harvesting, densitometric analysis indicated a Tf AA10A yield of 779 ± 40 µg L −1 in the culture medium without further optimization (Additional file 1: Figure S4). Although the lack of quantification data in other cyanobacterial secretion studies hinders widespread comparisons, to our knowledge, this is the highest yield reported to date for the secretion of a heterologous enzyme in cyanobacteria [24, 25, 63]. Although our reported yield of secreted LPMO is approximately 25% of previously reported yields for E. coli Tf AA10A expression (3 mg L −1 ), the majority of Tf AA10A in E. coli was exported to the periplasm [30], whereas the S. elongatus UTEX 2973 Tf AA10A was fully secreted to the medium.…”

Section: Discussionmentioning

confidence: 99%

“…Although protein export and secretion pathways in cyanobacteria have been studied for almost two decades, they remain poorly understood (see recent reviews [21, 23]). Proof-of-concept studies have shown that heterologous protein secretion in cyanobacteria is possible, however, these studies have generally utilized native signal peptides fused to reporter proteins and have suffered from low yields [24, 25]. In this work, we engineered an industrially relevant extracellular enzyme, a lytic polysaccharide monooxygenase (LPMO), into the fast-growing cyanobacterium S. elongatus UTEX 2973 to assess its secretion capabilities.…”

Section: Introductionmentioning

confidence: 99%

Expression and secretion of a lytic polysaccharide monooxygenase by a fast-growing cyanobacterium

Russo

Zedler

Wittmann

et al. 2019

Biotechnol Biofuels

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Background Cyanobacteria have the potential to become next-generation cell factories due to their ability to use CO 2 , light and inorganic nutrients to produce a range of biomolecules of commercial interest. Synechococcus elongatus UTEX 2973, in particular, is a fast-growing, genetically tractable, cyanobacterium that has garnered attention as a potential biotechnological chassis. To establish this unique strain as a host for heterologous protein production, we aimed to demonstrate expression and secretion of the industrially relevant Tf AA10A, a lytic polysaccharide monooxygenase from the Gram-positive bacterium Thermobifida fusca. Results Two variations of Tf AA10A were successfully expressed in S. elongatus UTEX 2973: One containing the native N-terminal, Sec-targeted, signal peptide and a second with a Tat-targeted signal peptide from the Escherichia coli trimethylamine- N -oxide reductase (TorA). Although the TorA signal peptide correctly targeted the protein to the plasma membrane, the majority of the TorA- Tf AA10A was found unprocessed in the plasma membrane with a small fraction of the mature protein ultimately translocated to the periplasm. The native Sec signal peptide allowed for efficient secretion of Tf AA10A into the medium with virtually no protein being found in the cytosol, plasma membrane or periplasm. Tf AA10A was demonstrated to be correctly cleaved and active on the model substrate phosphoric acid swollen cellulose. Additionally, expression and secretion only had a minor impact on cell growth. The secretion yield was estimated at 779 ± 40 µg L −1 based on densitometric analysis. To our knowledge, this is the highest secretion yield ever registered in cyanobacteria. Conclusions We have shown for the first time high-titer expression and secretion of an industrially relevant and catalytically active enzyme in S. elongatus UTEX 2973. This proof-of-concept study will be valuable for the development of novel and sustainable applications in the fields of bioremediation and biocatalysis. Electronic supplementary material The online version of this article (10.1186/s13068-019-1416-9) contains supplementary material, which is available to authorized users.

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Cyanobacterial leader peptides for protein secretion

Cited by 15 publications

References 27 publications

Novel adaptive responses revealed by transcription profiling of a Synechocystis sp. PCC 6803 ΔisiA mutant in the presence and absence of hydrogen peroxide

Novel adaptive responses revealed by transcription profiling of a Synechocystis sp. PCC 6803 ΔisiA mutant in the presence and absence of hydrogen peroxide

Secretome analysis of Anabaena sp. PCC 7120 and the involvement of the TolC‐homologue HgdD in protein secretion

Expression and secretion of a lytic polysaccharide monooxygenase by a fast-growing cyanobacterium

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