Effect of Signal Peptide on Stability and Folding of Escherichia coli Thioredoxin

Singh, Pranveer; Sharma, Likhesh; Kulothungan, S.; Adkar, Bharat V.; Prajapati, Ravindra Singh; Ali, P. Shaik Syed; Krishnan, Beena; Varadarajan, Raghavan

doi:10.1371/journal.pone.0063442

Cited by 67 publications

(48 citation statements)

References 61 publications

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“…These results indicated that the native signal sequence of Est1 interfered with correct protein folding, when not cleaved correctly during secretion. Similar observations for E. coli were made previously [25].…”

Section: Fig 1 (A) Expression Of Est1 With E Coli Bl21 (De3)star Pesupporting

confidence: 89%

Heterologous production of a feruloyl esterase from Pleurotus sapidus synthesizing feruloyl‐saccharide esters

Kelle

Nieter

Krings

et al. 2015

Biotech and App Biochem

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The feruloyl esterase (FAE) gene EST1 from the basidiomycete Pleurotus sapidus was heterologously expressed in Escherichia coli and Pichia pastoris. Catalytically active recombinant Est1 was secreted using P. pastoris as a host. For expression in P. pastoris, the expression vector pPIC9K was applied. The EST1 gene was cloned with an N-terminal α-mating factor pre-pro sequence and expressed under the control of a methanol inducible alcohol oxidase 1 promotor. Est1 was purified to homogeneity using ion exchange and hydrophobic interaction chromatography. The recombinant Est1 showed optima at pH 5.0 and 50 °C, and released ferulic acid from saccharide esters and from the natural substrate destarched wheat bran. Substrate specificity profile and descriptor-based analysis demonstrated unique properties, showing that Est1 did not fit into the current FAE classification model. Transferuloylation synthesis of feruloyl-saccharide esters was proven for mono- and disaccharides.

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Section: Fig 1 (A) Expression Of Est1 With E Coli Bl21 (De3)star Pesupporting

confidence: 89%

Heterologous production of a feruloyl esterase from Pleurotus sapidus synthesizing feruloyl‐saccharide esters

Kelle

Nieter

Krings

et al. 2015

Biotech and App Biochem

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“…17 Secretory proteins such as CCL11 only become functional after crossing the membrane and arriving to the appropriate cellular compartment and, consequently, being cleaved by signal peptidases. 49,50 The signal peptide plays important roles in targeting and membrane insertion, and after being cleaved can also exert other functions such as protecting cells from being killed by other cells. [49][50][51][52] Before cleavage, the signal peptide may have also important functions in protein folding and maturation.…”

Section: Resultsmentioning

confidence: 99%

“…49,50 The signal peptide plays important roles in targeting and membrane insertion, and after being cleaved can also exert other functions such as protecting cells from being killed by other cells. [49][50][51][52] Before cleavage, the signal peptide may have also important functions in protein folding and maturation. 52 Therefore, mutations in the signal peptide may interfere with such functions.…”

Section: Resultsmentioning

confidence: 99%

Evolution of CCL11: genetic characterization in lagomorphs and evidence of positive and purifying selection in mammals

2016

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The interactions between chemokines and their receptors are crucial for differentiation and activation of inflammatory cells. CC chemokine ligand 11 (CCL11) binds to CCR3 and to CCR5 that in leporids underwent gene conversion with CCR2. Here, we genetically characterized CCL11 in lagomorphs (leporids and pikas). All lagomorphs have a potentially functional CCL11, and the Pygmy rabbit has a mutation in the stop codon that leads to a longer protein. Other mammals also have mutations at the stop codon that result in proteins with different lengths. By employing maximum likelihood methods, we observed that, in mammals, CCL11 exhibits both signatures of purifying and positive selection. Signatures of purifying selection were detected in sites important for receptor binding and activation. Of the three sites detected as under positive selection, two were located close to the stop codon. Our results suggest that CCL11 is functional in all lagomorphs, and that the signatures of purifying and positive selection in mammalian CCL11 probably reflect the protein's biological roles.

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“…S3 in the supplemental material). The mechanism of protein secretion in eukaryotes and prokaryotes requires the signal sequence to facilitate transport of the translated protein to the membrane (31), and it also helps the protein attain the right conformation required to maintain its functionality (32). The fact that the major sequence difference between the FAD-I of ATCC 10953 and those of the other two strains is in the signal peptide domain of FAD-I ( Fig.…”

Section: Discussionmentioning

confidence: 99%

FAD-I, a Fusobacterium nucleatum Cell Wall-Associated Diacylated Lipoprotein That Mediates Human Beta Defensin 2 Induction through Toll-Like Receptor-1/2 (TLR-1/2) and TLR-2/6

Bhattacharyya¹,

Ghosh²,

Shokeen

et al. 2016

Infect Immun

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We previously identified a cell wall-associated protein from Fusobacterium nucleatum, a Gram-negative bacterium of the oral cavity, that induces human beta defensin 2 (hBD-2) in primary human oral epithelial cells (HOECs) and designated it FAD-I (Fusobacterium-associated defensin inducer). Here, we report differential induction of hBD-2 by different strains of F. nucleatum; ATCC 25586 and ATCC 23726 induce significantly more hBD-2 mRNA than ATCC 10953. Heterologous expression of plasmid-borne fadI from the highly hBD-2-inducing strains in a ⌬fadI mutant of ATCC 10953 resulted in hBD-2 induction to levels comparable to those of the highly inducing strains, indicating that FAD-I is the principal F. nucleatum agent for hBD-2 induction in HOECs. Moreover, anti-FAD-I antibodies blocked F. nucleatum induction of hBD-2 by more than 80%. Recombinant FAD-I (rFAD-I) expressed in Escherichia coli triggered levels of hBD-2 transcription and peptide release in HOECs similar to those of native FAD-I (nFAD-I) isolated from F. nucleatum ATCC 25586. Tandem mass spectrometry revealed a diacylglycerol modification at the cysteine residue in position 16 for both nFAD-I and rFAD-I. Cysteine-to-alanine substitution abrogated FAD-I's ability to induce hBD-2. Finally, FAD-I activation of hBD-2 expression was mediated via both Toll-like receptor-1/2 (TLR-1/2) and TLR-2/6 heterodimerization. Microbial molecules like FAD-I may be utilized in novel therapeutic ways to bolster the host innate immune response at mucosal surfaces.T he epithelial surfaces of the oral cavity are sites of active bacterial colonization. While colonizing, certain bacteria promote activation of human beta defensin (hBD) expression in the oral mucosa (1-3). By virtue of their antimicrobial and immunoregulatory properties, these epithelial-cell-derived innate response peptides contribute to the homeostasis between the bacterium and the host (4). Human beta defensin 2 (hBD-2) and hBD-3 are the two inducible members of the hBD peptide family that we and others have described in the oral cavity (1, 5-10). Interestingly, while hBD-3 is associated with the highly proliferating, nondifferentiated stratum basale of the oral mucosa, hBD-2 is compartmentalized in the more superficial stratum spinosum and stratum granulosum; i.e., nonproliferating yet differentiating regions of the oral mucosa (11,12). This, along with other results showing that hBD-2 is induced as a result of inflammation via MAPK or NF〉 (5) while hBD-3 is activated through epidermal growth factor receptor (13, 14), strongly suggests that the latter is more involved in wound healing while the former plays a more active role in inhibiting microbial invasion during mucosal-barrier disruption (15-17). Moreover, in addition to their antimicrobial properties (18, 19), both peptides have been shown to act as chemokines in recruiting lymphoid and myeloid cells from the bloodstream (20).Fusobacterium nucleatum, a ubiquitous Gram-negative bacterium of the human oral cavity, has been extensively studied for its propert...

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Effect of Signal Peptide on Stability and Folding of Escherichia coli Thioredoxin

Cited by 67 publications

References 61 publications

Heterologous production of a feruloyl esterase from Pleurotus sapidus synthesizing feruloyl‐saccharide esters

Heterologous production of a feruloyl esterase from Pleurotus sapidus synthesizing feruloyl‐saccharide esters

Evolution of CCL11: genetic characterization in lagomorphs and evidence of positive and purifying selection in mammals

FAD-I, a Fusobacterium nucleatum Cell Wall-Associated Diacylated Lipoprotein That Mediates Human Beta Defensin 2 Induction through Toll-Like Receptor-1/2 (TLR-1/2) and TLR-2/6

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